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Constantine A Stratakis - One of the best experts on this subject based on the ideXlab platform.

  • Variants in PRKAR1B cause a neurodevelopmental disorder with autism spectrum disorder, apraxia, and insensitivity to pain
    Genetics in Medicine, 2021
    Co-Authors: Felix Marbach, Constantine A Stratakis, Nikolaos Settas, Edra London, Georgi Stoyanov, Florian Erger, Jill A. Rosenfeld, Erin Torti, Chad Haldeman-englert, Evgenia Sklirou
    Abstract:

    Purpose We characterize the clinical and molecular phenotypes of six unrelated individuals with intellectual disability and autism spectrum disorder who carry heterozygous missense variants of the PRKAR1B gene, which encodes the R1β subunit of the cyclic AMP-dependent protein kinase A (PKA). Methods Variants of PRKAR1B were identified by single- or trio-exome analysis. We contacted the families and physicians of the six individuals to collect phenotypic information, performed in vitro analyses of the identified PRKAR1B -variants, and investigated PRKAR1B expression during embryonic development. Results Recent studies of large patient cohorts with neurodevelopmental disorders found significant enrichment of de novo missense variants in PRKAR1B . In our cohort, de novo origin of the PRKAR1B variants could be confirmed in five of six individuals, and four carried the same heterozygous de novo variant c.1003C>T (p.Arg335Trp; NM_001164760). Global developmental delay, autism spectrum disorder, and apraxia/dyspraxia have been reported in all six, and reduced pain sensitivity was found in three individuals carrying the c.1003C>T variant. PRKAR1B expression in the brain was demonstrated during human embryonal development. Additionally, in vitro analyses revealed altered basal PKA activity in cells transfected with variant-harboring PRKAR1B expression constructs. Conclusion Our study provides strong evidence for a PRKAR1B -related neurodevelopmental disorder.

  • First Somatic PRKAR1A Defect Associated With Mosaicism for Another PRKAR1A Mutation in a Patient With Cushing Syndrome.
    Journal of the Endocrine Society, 2021
    Co-Authors: Crystal Kamilaris, Fabio R. Faucz, Victoria C. Andriessen, Naris Nilubol, Chyi-chia Richard Lee, Mark A. Ahlman, Fady Hannah-shmouni, Constantine A Stratakis
    Abstract:

    Context Primary pigmented nodular adrenocortical disease (PPNAD) is a rare cause of ACTH-independent Cushing syndrome (CS) associated mostly with Carney complex (CNC), a rare autosomal dominant multiple neoplasia syndrome. More than two-thirds of familial cases and approximately one-third of sporadic cases of CNC harbor germline inactivating PRKAR1A defects. Increasingly sensitive technologies for the detection of genetic defects such as next-generation sequencing (NGS) have further highlighted the importance of mosaicism in human disease. Case Description A 33-year-old woman was diagnosed with ACTH-independent CS with abdominal computed tomography showing bilateral micronodular adrenal hyperplasia with a left adrenal adenoma. She underwent left adrenalectomy with pathology demonstrating PPNAD with a 1.5-cm pigmented adenoma. DNA analysis by Sanger sequencing revealed 2 different PRKAR1A variants in the adenoma that were absent from DNA extracted from blood and saliva: c.682C > T and c.974-2A > G. "Deep" NGS revealed that 0.31% of DNA copies extracted from blood and saliva did in fact carry the c.682C > T variant, suggesting low-level mosaicism for this defect. Conclusions We present a case of PPNAD due to low-level mosaicism for a PRKAR1A defect which led to the formation of an adenoma due to a second, adrenal-specific, somatic PRKAR1A mutation. The identification of mosaicism for PRKAR1A, depending on the number and distribution of cells affected has implications for genetic counseling and tumor surveillance. This is the first recorded case of a patient with PRKAR1A mosaicism, PPNAD, and an adenoma forming due to complete inactivation of PRKAR1A in adrenal tissue from a second, somatic-only, PRKAR1A coding sequence mutation.

  • c kit oncogene expression in PRKAR1A mutant adrenal cortex
    Endocrine-related Cancer, 2020
    Co-Authors: Kiran S. Nadella, Fabio R. Faucz, Constantine A Stratakis
    Abstract:

    Protein kinase A (PKA) regulatory subunit type 1A (PRKAR1A) defects lead to primary pigmented nodular adrenocortical disease (PPNAD). The KIT protooncogene (c-KIT) is not known to be expressed in the normal adrenal cortex (AC). In this study, we investigated the expression of c-KIT and its ligand, stem cell factor (SCF), in PPNAD and other cortisol-producing tumors of the adrenal cortex. mRNA and protein expression, by qRT-PCR, immunohistochemistry (IHC) and immunoblotting (IB), respectively, were studied. We then tested c-KIT and SCF responses to PRKAR1A introduction and PKA stimulation in adrenocortical cell lines CAR47 and H295R, which were also treated with the KIT inhibitor, imatinib mesylate (IM). Mice xenografted with H295R cells were treated with IM. There was increased c-KIT mRNA expression in PPNAD; IHC showed KIT and SCF immunoreactivity within certain nodular areas in PPNAD. IB data was consistent with IHC and mRNA data. PRKAR1A-deficient CAR47 cells expressed c-KIT; this was enhanced by forskolin and lowered by PRKAR1A reintroduction. Knockdown of PKA's catalytic subunit (PRKACA) by siRNA reduced c-KIT levels. Treatment of the CAR47 cells with IM resulted in reduced cell viability, growth arrest, and apoptosis. Treatment with IM of mice xenografted with H295 cells inhibited further tumor growth. We conclude that c-KIT is expressed in PPNAD, an expression that appears to be dependent on PRKAR1A and/or PKA activity. In a human adrenocortical cell line and its xenografts in mice, c-KIT inhibition decreased growth, suggesting that c-KIT inhibitors may be a reasonable alternative therapy to be tested in PPNAD, when other treatments are not optimal.

  • PRKAR1A haploinsufficiency ameliorates the growth hormone excess phenotype in Aip-deficient mice
    Human molecular genetics, 2020
    Co-Authors: Marie Helene Schernthaner-reiter, Matthew F. Starost, Giampaolo Trivellin, Thomas Roetzer, Johannes A. Hainfellner, Constantine A Stratakis
    Abstract:

    Mutations of the regulatory subunit (PRKAR1A) of the cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA), leading to activation of the PKA pathway, are the genetic cause of Carney complex which is frequently accompanied by somatotroph tumors. Aryl hydrocarbon receptor-interacting protein (AIP) mutations lead to somatotroph tumorigenesis in mice and humans. The mechanisms of AIP-dependent pituitary tumorigenesis are still under investigation and evidence points to a connection between the AIP and PKA pathways. In this study, we explore the combined effects of Aip and PRKAR1A deficiency on mouse phenotype and, specifically, pituitary histopathology. Aip+/- mice were compared with double heterozygous Aip+/-, PRKAR1A+/- mice. The phenotype (including histopathology and serological studies) was recorded at 3, 6, 9 and 12 months of age. Detailed pituitary histological and immunohistochemical studies were performed at 12 months. Twelve-month old Aip+/- mice demonstrated phenotypic and biochemical evidence of GH excess including significantly elevated insulin-like growth factor 1 levels, larger weight and body length, higher hemoglobin and cholesterol levels and a higher frequency of growth plate thickening in comparison to Aip+/, PRKAR1A+/- mice. Pituitary histopathology did not uncover any pituitary adenomas or somatotroph hyperplasia in either group. These results demonstrate a slow progression from elevated GH release to the formation of overt somatotropinomas in Aip+/- mice; the acromegalic phenotype of these mice is surprisingly ameliorated in Aip+/-, PRKAR1A+/- mice. This highlights the complexities of interaction between the AIP and PKA pathway. Specifically targeting GH secretion rather than somatotroph proliferation may be an advantage in the medical treatment of AIP-dependent human acromegaly.

  • Abstract MP166: PRKAR1A Deficiency Abrogates Cardiac Hypertrophy Through Inhibition of Mitochondrial Fission
    Circulation Research, 2020
    Co-Authors: Yuening Liu, Constantine A Stratakis, Peng Xia, Jingrui Chen, Patricia W Bandettini, Lawrence S. Kirschner, Zhaokang Cheng
    Abstract:

    Protein kinase A (PKA) is pivotal for cardiac function of human heart, and its dysregulation is involved with various cardiac pathologies. PKA regulatory subunit 1α (R1α, encoded by PRKAR1A gene) controls PKA kinase activity by sequestering the PKA catalytic subunits. Patients with PRKAR1A mutations are often diagnosed with Carney complex (CNC) and may die prematurely from cardiac complications such as heart failure. However, it remains unknown whether PRKAR1A deficiency interferes with normal heart growth during postnatal development. Here, we show that left ventricular mass is reduced in young CNC patients with PRKAR1A mutations or deletions. To investigate the impact of PRKAR1A deficiency on heart growth, we generated cardiac-specific PRKAR1A heterozygous knockout mice. Ablation of the PRKAR1A gene in mice increased cardiac PKA activity, reduced heart weight to body weight ratio and cardiomyocyte size without altering contractile function. Cardiomyocyte hypertrophy in response to activation of the α1-adrenergic receptor, was completely abolished by silencing of PRKAR1A . Mechanistically, depletion of PRKAR1A provoked PKA-dependent phosphorylation of the mitochondrial fission protein Drp1 at S637, resulting in impaired mitochondrial fission and diminished cardiomyocyte hypertrophy. In conclusion, PRKAR1A deficiency abrogates cardiac hypertrophy during postnatal development, likely through inhibiting Drp1-mediated mitochondrial fission. Our study provides novel mechanistic insights regarding the cardiac mortality associated with CNC.

Jerome Bertherat - One of the best experts on this subject based on the ideXlab platform.

  • ARMC5 variants in PRKAR1A-mutated patients modify cortisol levels and Cushing's syndrome.
    Endocrine-related cancer, 2020
    Co-Authors: Andrea Gutierrez Maria, Fabio R. Faucz, Ludivine Drougat, Nikolaos Settas, Jerome Bertherat, Fady Hannah-shmouni, Christina Tatsi, Annabel Berthon, Constantine A Stratakis
    Abstract:

    Mutations in the protein kinase A (PKA) regulatory subunit type 1A (PRKAR1A) and armadillo repeat-containing 5 (ARMC5) genes cause Cushing's syndrome (CS) due to primary pigmented nodular adrenocortical disease (PPNAD) and primary bilateral macronodular adrenocortical hyperplasia (PBMAH), respectively. Between the two genes, ARMC5 is highly polymorphic with several variants in the population, whereas PRKAR1A has very little, if any, non-pathogenic variation in its coding sequence. We tested the hypothesis that ARMC5 variants may affect the clinical presentation of PPNAD and CS among patients with PRKAR1A mutations. In this study, 91 patients with PPNAD due to PRKAR1A mutations were tested for abnormal cortisol secretion or CS and for ARMC5 sequence variants. Abnormal cortisol secretion was present in 71 of 74 patients with ARMC5 variants, whereas 11 of 17 patients negative for ARMC5 variants did not have hypercortisolemia. The presence of ARMC5 variants was a statistically strong predictor of CS among patients with PRKAR1A mutations (P < 0.001). Among patients with CS due to PPNAD, ARMC5 variants were associated with lower cortisol levels at baseline (P = 0.04) and after high dose dexamethasone administration (P = 0.02). The ARMC5 p.I170V variant increased ARMC5 protein accumulation in vitro and decreased viability of NCI-H295 cells (but not HEK 293T cells). PPNAD tissues with ARMC5 variants showed stronger ARMC5 protein expression than those that carried a normal ARMC5 sequence. Taken together, our results suggest that ARMC5 variants among patients with PPNAD due to PRKAR1A defects may play the role of a genetic modifier for the presence and severity of hypercortisolemia.

  • functional characterization of PRKAR1A mutations reveals a unique molecular mechanism causing acrodysostosis but multiple mechanisms causing carney complex
    Journal of Biological Chemistry, 2015
    Co-Authors: Yara Rhayem, Agnes Linglart, Jerome Bertherat, Catherine Le Stunff, Waed Abdel Khalek, Colette Auzan, Alain Couvineau, Caroline Silve, Eric Clauser
    Abstract:

    The main target of cAMP is PKA, the main regulatory subunit of which (PRKAR1A) presents mutations in two genetic disorders: acrodysostosis and Carney complex. In addition to the initial recurrent mutation (R368X) of the PRKAR1A gene, several missense and nonsense mutations have been observed recently in acrodysostosis with hormonal resistance. These mutations are located in one of the two cAMP-binding domains of the protein, and their functional characterization is presented here. Expression of each of the PRKAR1A mutants results in a reduction of forskolin-induced PKA activation (measured by a reporter assay) and an impaired ability of cAMP to dissociate PRKAR1A from the catalytic PKA subunits by BRET assay. Modeling studies and sensitivity to cAMP analogs specific for domain A (8-piperidinoadenosine 3',5'-cyclic monophosphate) or domain B (8-(6-aminohexyl)aminoadenosine-3',5'-cyclic monophosphate) indicate that the mutations impair cAMP binding locally in the domain containing the mutation. Interestingly, two of these mutations affect amino acids for which alternative amino acid substitutions have been reported to cause the Carney complex phenotype. To decipher the molecular mechanism through which homologous substitutions can produce such strikingly different clinical phenotypes, we studied these mutations using the same approaches. Interestingly, the Carney mutants also demonstrated resistance to cAMP, but they expressed additional functional defects, including accelerated PRKAR1A protein degradation. These data demonstrate that a cAMP binding defect is the common molecular mechanism for resistance of PKA activation in acrodysosotosis and that several distinct mechanisms lead to constitutive PKA activation in Carney complex.

  • Comparison of the effects of PRKAR1A and PRKAR2B depletion on signaling pathways, cell growth, and cell cycle control of adrenocortical cells.
    Hormone and metabolic research = Hormon- und Stoffwechselforschung = Hormones et metabolisme, 2014
    Co-Authors: F. Basso, Maria Nesterova, Constantine A Stratakis, Jerome Bertherat, Bruno Ragazzon, F. Rocchetti, Stephanie Rodriguez, F. Cormier, Marthe Rizk-rabin
    Abstract:

    The cyclic AMP/protein kinase A signaling cascade is one of the main pathways involved in the pathogenesis of adrenocortical tumors. The PKA R1A and R2B proteins are the most abundant regulatory subunits in endocrine tissues. Inactivating mutations of PRKAR1A are associated with Carney complex and a subset of sporadic tumors and the abundance of R2B protein is low in a subset of secreting adrenocortical adenomas. We previously showed that PRKAR1A and PRKAR2B inactivation have anti-apoptotic effects on the adrenocortical carcinoma cell line H295R. The aim of this study was to compare the effects of PRKAR1A and PRKAR2B depletion on cell proliferation, apoptosis, cell signaling pathways, and cell cycle regulation. We found that PRKAR2B depletion is compensated by an upregulation of R1A protein, whereas PRKAR1A depletion has no effect on the production of R2B. The depletion of either PRKAR1A or PRKAR2B promotes the expression of Bcl-xL and resistance to apoptosis; and is associated with a high percentage of cells in S and G2 phase, activates PKA and MEK/ERK pathways, and impairs the expression of IkB leading to activate the NF-κB pathway. However, we observed differences in the regulation of cyclins. The depletion of PRKAR1A leads to the accumulation of cyclin D1 and p27kip, whereas the depletion of PRKAR2B promotes the accumulation of cyclin A, B, cdk1, cdc2, and p21Cip. In conclusion, although the depletion of PRKAR1A and PRKAR2B in adrenocortical cells has similar effects on cell proliferation and apoptosis; loss of these PKA subunits differentially affects cyclin expression.

  • Protein kinase A alterations in endocrine tumors.
    Hormone and Metabolic Research, 2012
    Co-Authors: Bo Yu, Marthe Rizk-rabin, Bruno Ragazzon, Jerome Bertherat
    Abstract:

    Various molecular and cellular alterations of the cyclic adenosine monophosphate (cAMP) pathway have been observed in endocrine tumors. Since protein kinase A (PKA) is a central key component of the cAMP pathway, studies of the alterations of PKA subunits in endocrine tumors reveal new aspects of the mechanisms of cAMP pathway alterations in human diseases. So far, most alterations have been observed for the regulatory subunits, mainly PRKAR1A and to a lower extent, PRKAR2B . One of the best examples of such alteration today is the multiple neoplasia syndrome Carney complex (CNC). The most common endocrine gland manifestations of CNC are pituitary GH-secreting adenomas, thyroid tumors, testicular tumors, and ACTH-independent Cushing’s syndrome due to primary pigmented nodular adrenocortical disease (PPNAD). Heterozygous germline inactivating mutations of the PKA regulatory subunit RIα gene ( PRKAR1A ) are observed in about two-third of CNC patients, and also in patients with isolated PPNAD. PRKAR1A is considered as a tumor suppressor gene. Interestingly, these mutations can also be observed as somatic alterations in sporadic endocrine tumors. More than 120 different PRKAR1A mutations have been found today. Most of them lead to an unstable mutant mRNA, which will be degraded by nonsense mediated mRNA decay. In vitro and in vivo functional studies are in progress to understand the mechanisms of endocrine tumor development due to PKA regulatory subunits inactivation. PRKAR1A mutations stimulate in most models PKA activity, mimicking in some way cAMP pathway constitutive activation. Cross-talks with other signaling pathways summarized in this review have been described and might participate in endocrine tumorigenesis.

  • mutations and polymorphisms in the gene encoding regulatory subunit type 1 alpha of protein kinase a PRKAR1A an update
    Human Mutation, 2010
    Co-Authors: Anelia Horvath, Kit Man Tsang, Lionel Groussin, Jerome Bertherat, Laure Cazabat, R Libe, Marine Guillaudbataille
    Abstract:

    PRKAR1A encodes the regulatory subunit type 1-alpha (RIalpha) of the cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA). Inactivating PRKAR1A mutations are known to be responsible for the multiple neoplasia and lentiginosis syndrome Carney complex (CNC). To date, at least 117 pathogenic variants in PRKAR1A have been identified (online database: http://PRKAR1A.nichd.nih.gov). The majority are subject to nonsense mediated mRNA decay (NMD), leading to RIalpha haploinsufficiency and, as a result, activated cAMP signaling. Recently, it became apparent that CNC may be caused not only by RIalpha haploinsufficiency, but also by the expression of altered RIalpha protein, as proven by analysis of expressed mutations in the gene, consisting of amino acid substitutions and in-frame genetic alterations. In addition, a new subgroup of mutations that potentially escape NMD and result in CNC through altered (rather than missing) protein has been analyzed-these are frame-shifts in the 3' end of the coding sequence that shift the stop codon downstream of the normal one. The mutation detection rate in CNC patients is recently estimated at above 60%; PRKAR1A mutation-negative CNC patients are characterized by significant phenotypic heterogeneity. In this report, we present a comprehensive analysis of all presently known PRKAR1A sequence variations and discuss their molecular context and clinical phenotype.

Lawrence S. Kirschner - One of the best experts on this subject based on the ideXlab platform.

  • Abstract MP166: PRKAR1A Deficiency Abrogates Cardiac Hypertrophy Through Inhibition of Mitochondrial Fission
    Circulation Research, 2020
    Co-Authors: Yuening Liu, Constantine A Stratakis, Peng Xia, Jingrui Chen, Patricia W Bandettini, Lawrence S. Kirschner, Zhaokang Cheng
    Abstract:

    Protein kinase A (PKA) is pivotal for cardiac function of human heart, and its dysregulation is involved with various cardiac pathologies. PKA regulatory subunit 1α (R1α, encoded by PRKAR1A gene) controls PKA kinase activity by sequestering the PKA catalytic subunits. Patients with PRKAR1A mutations are often diagnosed with Carney complex (CNC) and may die prematurely from cardiac complications such as heart failure. However, it remains unknown whether PRKAR1A deficiency interferes with normal heart growth during postnatal development. Here, we show that left ventricular mass is reduced in young CNC patients with PRKAR1A mutations or deletions. To investigate the impact of PRKAR1A deficiency on heart growth, we generated cardiac-specific PRKAR1A heterozygous knockout mice. Ablation of the PRKAR1A gene in mice increased cardiac PKA activity, reduced heart weight to body weight ratio and cardiomyocyte size without altering contractile function. Cardiomyocyte hypertrophy in response to activation of the α1-adrenergic receptor, was completely abolished by silencing of PRKAR1A . Mechanistically, depletion of PRKAR1A provoked PKA-dependent phosphorylation of the mitochondrial fission protein Drp1 at S637, resulting in impaired mitochondrial fission and diminished cardiomyocyte hypertrophy. In conclusion, PRKAR1A deficiency abrogates cardiac hypertrophy during postnatal development, likely through inhibiting Drp1-mediated mitochondrial fission. Our study provides novel mechanistic insights regarding the cardiac mortality associated with CNC.

  • PRKAR1A deficiency impedes hypertrophy and reduces heart size.
    Physiological reports, 2020
    Co-Authors: Yuening Liu, Constantine A Stratakis, Peng Xia, Jingrui Chen, Lawrence S. Kirschner, W. Patricia Bandettini, Zhaokang Cheng
    Abstract:

    Protein kinase A (PKA) activity is pivotal for proper functioning of the human heart, and its dysregulation has been implicated in a variety of cardiac pathologies. PKA regulatory subunit 1α (R1α, encoded by the PRKAR1A gene) is highly expressed in the heart, and controls PKA kinase activity by sequestering PKA catalytic subunits. Patients with PRKAR1A mutations are often diagnosed with Carney complex (CNC) in early adulthood, and may die later in life from cardiac complications such as heart failure. However, it remains unknown whether PRKAR1A deficiency interferes with normal heart development. Here, we showed that left ventricular mass was reduced in young CNC patients with PRKAR1A mutations or deletions. Cardiac-specific heterozygous ablation of PRKAR1A in mice increased cardiac PKA activity, and reduced heart weight and cardiomyocyte size without altering contractile function at 3 months of age. Silencing of PRKAR1A, or stimulation with the PKA activator forskolin completely abolished α1-adrenergic receptor-mediated cardiomyocyte hypertrophy. Mechanistically, depletion of PRKAR1A provoked PKA-dependent inactivating phosphorylation of Drp1 at S637, leading to impaired mitochondrial fission. Pharmacologic inhibition of Drp1 with Mdivi 1 diminished hypertrophic growth of cardiomyocytes. In conclusion, PRKAR1A deficiency suppresses cardiomyocyte hypertrophy and impedes heart growth, likely through inhibiting Drp1-mediated mitochondrial fission. These findings provide a potential novel mechanism for the cardiac manifestations associated with CNC.

  • knockdown of PRKAR1A the gene responsible for carney complex interferes with differentiation in osteoblastic cells
    Molecular Endocrinology, 2014
    Co-Authors: Mei Zhang, Parmeet Kaur Manchanda, Qianben Wang, Lawrence S. Kirschner
    Abstract:

    PRKAR1A is the gene encoding the type 1A regulatory subunit of protein kinase A, and it is the cause of the inherited human tumor syndrome Carney complex. Data from our laboratory has demonstrated that PRKAR1A loss causes tumors in multiple cell lineages, including neural crest cells and osteoblasts. We have proposed that one mechanism by which tumorigenesis occurs is through the failure of terminal differentiation. In the present study, we directly test the effects of PRKAR1A reduction on osteogenic differentiation in mouse and human cells in vitro. We found that PRKAR1A levels noticeably increased during osteoblastic differentiation, indicating a positive correlation between the expression of PRKAR1A and osteogenic potential. To validate this hypothesis, we generated stable PRKAR1A knockdown in both mouse and human cells. These cells displayed significantly suppressed bone nodule formation and decreased expression of osteoblast markers such as osteocalcin and osteopontin. These observations imply that the antiosteogenic effect of PRKAR1A ablation is not species or cell line specific. Furthermore, because Runt-related transcription factor-2 (Runx2) is a key mediator of osteoblast differentiation, we reasoned that the function of this transcription factor may be inhibited by PRKAR1A knockdown. Chromatin immunoprecipitation and luciferase assays demonstrated that PRKAR1A ablation repressed DNA binding and function of Runx2 at its target genes. Additionally, we determined that this effect is likely due to reductions in the Runx2-cooperating transcription factors forkhead box O1 and activating transcription factor 4. Taken together, this study provides direct evidence that ablation of PRKAR1A interferes with signaling pathways necessary for osteoblast differentiation.

  • The Protein Kinase A Regulatory Subunit R1A (PRKAR1A) Plays Critical Roles in Peripheral Nerve Development
    The Journal of neuroscience : the official journal of the Society for Neuroscience, 2013
    Co-Authors: Li Guo, Audrey A. Lee, Tilat A. Rizvi, Nancy Ratner, Lawrence S. Kirschner
    Abstract:

    Signaling through cAMP has been implicated in Schwann cell (SC) proliferation and myelination, but the signaling pathway components downstream of cAMP required for SC function remain unknown. Protein kinase A (PKA) is a potential downstream effector of cAMP. Here, we induced loss of PRKAR1A, the gene encoding the type 1A regulatory subunit of PKA, in SC to study its role in nerve development; loss of PRKAR1A is predicted to elevate PKA activity. Conditional PRKAR1A knock-out in mouse SC (PRKAR1A-SCKO) resulted in a dramatic and persistent axonal sorting defect, and unexpectedly decreased SC proliferation in PRKAR1A-SCKO nerves in vivo. Effects were cell autonomous as they were recapitulated in vitro in PRKAR1A-SCKO SC, which showed elevated PKA activity. In the few SCs sorted into 1:1 relationships with axons in vivo, SC myelination was premature in PRKAR1A-SCKO nerves, correlating with global increase in the cAMP-regulated transcription factor Oct-6 and expression of myelin basic protein. These data reveal a previously unknown role of PKA in axon sorting, an unexpected inhibitory role of PKA on SC cell proliferation in vivo and define the importance of PRKAR1A in peripheral nerve development.

  • PRKAR1A and the evolution of pituitary tumors.
    Molecular and Cellular Endocrinology, 2010
    Co-Authors: Lawrence S. Kirschner
    Abstract:

    Carney complex (CNC) is an inherited tumor predisposition associated with pituitary tumors, including GH-producing pituitary adenomas and rare reports of prolactinomas. This disease is caused by mutations in PRKAR1A, which encodes the type 1A regulatory subunit of the cAMP-dependent protein kinase, PKA. Loss of PRKAR1A causes enhanced PKA signaling, which leads to pituitary tumorigenesis. Mutations in the gene have not been detected in sporadic pituitary tumors, but there is some data to suggest that non-genomic mechanisms may cause loss of protein expression. Unlike CNC patients, mice heterozygous for PRKAR1A mutations do not develop pituitary tumors, although complete knockout of the gene in the Pit1 lineage of the pituitary produces GH-secreting pituitary adenomas. These data indicate that complete loss of PRKAR1A/PRKAR1A is able to cause pituitary tumors in mice and men. The pattern of tumors is likely related to the signaling pathways employed in specific pituitary cell types.

Lionel Groussin - One of the best experts on this subject based on the ideXlab platform.

  • mutations and polymorphisms in the gene encoding regulatory subunit type 1 alpha of protein kinase a PRKAR1A an update
    Human Mutation, 2010
    Co-Authors: Anelia Horvath, Kit Man Tsang, Lionel Groussin, Jerome Bertherat, Laure Cazabat, R Libe, Marine Guillaudbataille
    Abstract:

    PRKAR1A encodes the regulatory subunit type 1-alpha (RIalpha) of the cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA). Inactivating PRKAR1A mutations are known to be responsible for the multiple neoplasia and lentiginosis syndrome Carney complex (CNC). To date, at least 117 pathogenic variants in PRKAR1A have been identified (online database: http://PRKAR1A.nichd.nih.gov). The majority are subject to nonsense mediated mRNA decay (NMD), leading to RIalpha haploinsufficiency and, as a result, activated cAMP signaling. Recently, it became apparent that CNC may be caused not only by RIalpha haploinsufficiency, but also by the expression of altered RIalpha protein, as proven by analysis of expressed mutations in the gene, consisting of amino acid substitutions and in-frame genetic alterations. In addition, a new subgroup of mutations that potentially escape NMD and result in CNC through altered (rather than missing) protein has been analyzed-these are frame-shifts in the 3' end of the coding sequence that shift the stop codon downstream of the normal one. The mutation detection rate in CNC patients is recently estimated at above 60%; PRKAR1A mutation-negative CNC patients are characterized by significant phenotypic heterogeneity. In this report, we present a comprehensive analysis of all presently known PRKAR1A sequence variations and discuss their molecular context and clinical phenotype.

  • inactivation of the carney complex gene 1 protein kinase a regulatory subunit 1a inhibits smad3 expression and tgfβ stimulated apoptosis in adrenocortical cells
    Cancer Research, 2009
    Co-Authors: Bruno Ragazzon, Lionel Groussin, Laure Cazabat, Karine Perlemoine, Marthe Rizkrabin, Guillaume Assie, Helene Fierrard, Antoine Martinez, Jerome Bertherat
    Abstract:

    The cyclic AMP signaling pathway can be altered at multiple levels in endocrine tumors. Its central component is the protein kinase A (PKA). Carney complex (CNC) is a hereditary multiple neoplasia syndrome resulting from inactivating mutations of the gene encoding the PKA type I α regulatory subunit ( PRKAR1A ). Primary pigmented nodular adrenocortical disease is the most frequent endocrine tumor of CNC. Transforming growth factor β (TGFβ) regulates adrenal cortex physiology and signals through SMAD2/3. We used an interference approach to test the effects of PRKAR1A inactivation on PKA and TGFβ pathways and on apoptosis in adrenocortical cells. PRKAR1A silencing stimulates PKA activity and increases transcriptional activity of a PKA reporter construct and expression of the endogenous PKA target, NR4A2 , under basal conditions or after forskolin stimulation. PRKAR1A inactivation also decreased SMAD3 mRNA and protein levels via PKA, altering the cellular response to TGFβ. SMAD3 expression was also inhibited by adrenocorticorticotropic hormone in the mouse adrenal gland and by forskolin in H295R cells. TGFβ stimulates apoptosis in H295R cells, and this effect was counteracted by PRKAR1A inactivation. PRKAR1A silencing decreased the percentage of apoptotic cells and the cleavage of apoptosis mediators [caspase-3, poly(ADP-ribose) polymerase, and lamin A/C]. Inactivating mutations of PRKAR1A observed in adrenocortical tumors alter SMAD3, leading to resistance to TGFβ-induced apoptosis. This cross-talk between the PKA and the TGFβ signaling pathways reveals a new mechanism of endocrine tumorigenesis. [Cancer Res 2009;69(18):7278–84]

  • mutations in regulatory subunit type 1a of cyclic adenosine 5 monophosphate dependent protein kinase PRKAR1A phenotype analysis in 353 patients and 80 different genotypes
    The Journal of Clinical Endocrinology and Metabolism, 2009
    Co-Authors: Anelia Horvath, Lionel Groussin, Sosipatros Boikos, Jerome Bertherat, Sophie Grabar, Laure Cazabat, R Libe, F Renecorail
    Abstract:

    Background: The “complex of myxomas, spotty skin pigmentation, and endocrine overactivity,” or “Carney complex” (CNC), is caused by inactivating mutations of the regulatory subunit type 1A of the cAMP-dependent protein kinase (PRKAR1A) gene and as yet unknown defect(s) in other gene(s). Delineation of a genotype-phenotype correlation for CNC patients is essential for understanding PRKAR1A function and providing counseling and preventive care. Methods: A transatlantic consortium studied the molecular genotype and clinical phenotype of 353 patients (221 females and 132 males, age 34 ± 19 yr) who carried a germline PRKAR1A mutation or were diagnosed with CNC and/or primary pigmented nodular adrenocortical disease. Results: A total of 258 patients (73%) carried 80 different PRKAR1A mutations; 114 (62%) of the index cases had a PRKAR1A mutation. Most PRKAR1A mutations (82%) led to lack of detectable mutant protein (nonexpressed mutations) because of nonsense mRNA mediated decay. Patients with a PRKAR1A mutation were more likely to have pigmented skin lesions, myxomas, and thyroid and gonadal tumors; they also presented earlier with these tumors. Primary pigmented nodular adrenocortical disease occurred earlier, was more frequent in females, and was the only manifestation of CNC with a gender predilection. Mutations located in exons were more often associated with acromegaly, myxomas, lentigines, and schwannomas, whereas the frequent c.491-492delTG mutation was commonly associated with lentigines, cardiac myxomas, and thyroid tumors. Overall, nonexpressed PRKAR1A mutations were associated with less severe disease. Conclusion: CNC is genetically and clinically heterogeneous. Certain tumors are more frequent, with specific mutations providing some genotype-phenotype correlation for PRKAR1A mutations.

  • wnt β catenin and 3 5 cyclic adenosine 5 monophosphate protein kinase a signaling pathways alterations and somatic β catenin gene mutations in the progression of adrenocortical tumors
    The Journal of Clinical Endocrinology and Metabolism, 2008
    Co-Authors: Lionel Groussin, Bruno Ragazzon, Rossella Libé, Sebastien Gaujoux, Frederique Tissier, Pierre Launay, Anne Audebourg, B Dousset, Xavier Bertagna
    Abstract:

    Background: The Wnt/β-catenin and cAMP signaling pathways play an important role in adrenal cortex tumorigenesis. Somatic activating mutations of the β-catenin gene (CTNNB1) are the most frequent genetic defects identified both in adrenocortical adenomas (ACAs) and adrenocortical cancers (ACCs). PRKAR1A mutations leading to cAMP pathway dysregulation are observed in primary pigmented nodular adrenocortical diseases (PPNADs) and some sporadic ACAs. Objective: The objective of the investigation was to study Wnt/β-catenin dysregulation in adrenocortical tumors (ACTs) with cAMP pathway genetic alteration and search for secondary CTNNB1 somatic mutations in heterogeneous tumors. Patients and methods: Nine PPNADs, including five with macronodules, three ACAs with PRKAR1A somatic mutations, and one heterogeneous tumor with ACC developed within an ACA, were studied by immunohistochemistry and DNA sequencing. Results: β-Catenin accumulation was observed in all PPNADs, ACAs with PRKAR1A mutations, and the ACC compo...

  • Large deletions of the PRKAR1A gene in Carney complex.
    Clinical cancer research : an official journal of the American Association for Cancer Research, 2008
    Co-Authors: Anelia Horvath, Ioannis Bossis, Christoforos Giatzakis, Elizabeth Levine, Frank Weinberg, Elise Meoli, A. Robinson-white, Jennifer Siegel, Payal D. Soni, Lionel Groussin
    Abstract:

    Purpose: Since the identification of PRKAR1A mutations in Carney complex, substitutions and small insertions/deletions have been found in ∼70% of the patients. To date, no germ-line PRKAR1A deletion and/or insertion exceeded a few base pairs (up to 15). Although a few families map to chromosome 2, it is possible that current sequencing techniques do not detect larger gene changes in PRKAR1A –mutation-negative individuals with Carney complex. Experimental Design: To screen for gross alterations of the PRKAR1A gene, we applied Southern hybridization analysis on 36 unrelated Carney complex patients who did not have small intragenic mutations or large aberrations in PRKAR1A , including the probands from two kindreds mapping to chromosome 2. Results: We found large PRKAR1A deletions in the germ-line of two patients with Carney complex, both sporadic cases; no changes were identified in the remaining patients, including the two chromosome-2-mapping families. In the first patient, the deletion is expected to lead to decreased PRKAR1A mRNA levels but no other effects on the protein; the molecular phenotype is predicted to be PRKAR1A haploinsufficiency, consistent with the majority of PRKAR1A mutations causing Carney complex. In the second patient, the deletion led to in-frame elimination of exon 3 and the expression of a shorter protein, lacking the primary site for interaction with the catalytic protein kinase A subunit. In vitro transfection studies of the mutant PRKAR1A showed impaired ability to bind cyclic AMP and activation of the protein kinase A enzyme. The patient bearing this mutation had a more-severe-than-average Carney complex phenotype that included the relatively rare psammomatous melanotic schwannoma. Conclusions: Large PRKAR1A deletions may be responsible for Carney complex in patients that do not have PRKAR1A gene defects identifiable by sequencing. Preliminary data indicate that these patients may have a different phenotype especially if their defect results in an expressed, abnormal version of the PRKAR1A protein.

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  • deletions of the PRKAR1A locus at 17q24 2 q24 3 in carney complex genotype phenotype correlations and implications for genetic testing
    The Journal of Clinical Endocrinology and Metabolism, 2014
    Co-Authors: Paraskevi Salpea, Anelia Horvath, Fabio R. Faucz, Edra London, Annalisa Vetro, Isaac Levy, Evgenia Gourgari, Andrew Dauber, Ingrid A Holm, Patrick J Morrison
    Abstract:

    Background: Carney complex (CNC) is a multiple neoplasia syndrome caused by PRKAR1A-inactivating mutations. One-third of the patients, however, have no detectable PRKAR1A coding sequence defects. Small deletions of the gene were previously reported in few patients, but large deletions of the chromosomal PRKAR1A locus have not been studied systematically in a large cohort of patients with CNC. Setting: A tertiary care referral center was the setting for analysis of an international cohort of patients with CNC. Methods: Methods included genome-wide array analysis followed by fluorescent in situ hybridization, mRNA, and other studies as well as a retrospective analysis of clinical information and phenotype-genotype correlation. Results: We detected 17q24.2-q24.3 deletions of varying size that included the PRKAR1A gene in 11 CNC patients (of 51 tested). Quantitative PCR showed that these patients had significantly lower PRKAR1A mRNA levels. Phenotype varied but was generally severe and included manifestations...

  • eLS - Genetics of Carney Complex
    eLS, 2013
    Co-Authors: Anelia Horvath, Constantine A Stratakis
    Abstract:

    Carney complex (CNC) is an autosomal dominant multiple neoplasia and lentiginosis syndrome characterised by spotty skin lesions, cardiac and other myxomas and different types of endocrine tumours. The PRKAR1A gene, which codes for the regulatory subunit type 1-alpha of the cyclic adenosine monophosphate (cAMP)-activated protein kinase A, is responsible for more than two-thirds of the cases of CNC described to date. Currently, more than 120 different disease-causing PRKAR1A sequence variants have been reported. Other involved genes for adrenal hyperplasias include phosphodiesterases PDE11A and PDE8B. Additional genes are likely to be identified that may expand our understanding on the pathophysiology of the cAMP signalling pathway and how genetic defects cause CNC and its individual components such as adrenal tumours. Key Concepts: Inactivating PRKAR1A mutations cause CNC because PRKAR1A haploinsufficiency leads to uncontrolled PKA activity. The majority (∼80%) of PRKAR1A mutations causing CNC result in an early stop codon generation and degradation of the mutant RNA through nonsense-mediated RNA decay (NMD). Recently, novel types of PRKAR1A mutations have been described: large gene rearrangements and small indels resulting in an elongated protein through downstream shift of the stop codon. The penetrance of PRKAR1A mutations is more than 95% by the age of 50 years. PDE11A and PDE8B genetic defects contribute to the CNC phenotype, but they may also independently cause forms of adrenal hyperplasia, mainly isolated micronodular adrenocortical disease (iMAD). Genotype–phenotype correlation is limited, but there are certain mutations that cause more frequently certain endocrine manifestations such as isolated Cushing syndrome. The use of new genomic techniques has led to the identification of new genetic defects in PRKAR1A and related genes in CNC. Keywords: Carney complex; PRKAR1A; mutations; PDE11A; PDE8B; genotype-phenotype correlations

  • PRKAR1A gene analysis and protein kinase A activity in endometrial tumors.
    Endocrine-related cancer, 2012
    Co-Authors: A. Tsigginou, Maria Nesterova, Anelia Horvath, Sosipatros Boikos, Charalampos Lyssikatos, Eirini I. Bimpaki, C Papageorgiou, C Dimitrakakis, A Rodolakis, Constantine A Stratakis
    Abstract:

    PRKAR1A codes for the type 1a regulatory subunit (RIa) of the cAMP-dependent protein kinase A (PKA), an enzyme with an important role in cell cycle regulation and proliferation. PKA dysregulation has been found in various tumors, and PRKAR1A-inactivating mutations have been reported in mostly endocrine neoplasias. In this study, we investigated PKA activity and the PRKAR1A gene in normal and tumor endometrium. Specimens were collected from 31 patients with endometrial cancer. We used as controls 41 samples of endometrium that were collected from surrounding normal tissues or from women undergoing gynecological operations for other reasons. In all samples, we sequenced the PRKAR1A-coding sequence and studied PKA subunit expression; we also determined PKA activity and cAMP binding. PRKAR1A mutations were not found. However, PKA regulatory subunit protein levels, both RIa and those of regulatory subunit type 2b (RIIb), were lower in tumor samples; cAMP binding was also lower in tumors compared with normal endometrium (P!0.01). Free PKA activity was higher in tumor samples compared with that of control tissue (P!0.01). There are significant PKA enzymatic abnormalities in tumors of the endometrium compared with surrounding normal tissue; as these were not due to PRKAR1A mutations, other mechanisms affecting PKA function ought to be explored. Endocrine-Related Cancer (2012) 19 457‐462

  • activation of cyclic amp signaling leads to different pathway alterations in lesions of the adrenal cortex caused by germline PRKAR1A defects versus those due to somatic gnas mutations
    The Journal of Clinical Endocrinology and Metabolism, 2012
    Co-Authors: Madson Q Almeida, Michael T Collins, Anelia Horvath, Eirini I. Bimpaki, Monalisa F Azevedo, Paraskevi Xekouki, Lefkothea Karaviti, George S Jeha, Nisan Bhattacharyya, Chris Cheadle
    Abstract:

    Context: The overwhelming majority of benign lesions of the adrenal cortex leading to Cushing syndrome are linked to one or another abnormality of the cAMP or protein kinase pathway. PRKAR1A-inactivating mutations are responsible for primary pigmented nodular adrenocortical disease, whereas somatic GNAS activating mutations cause macronodular disease in the context of McCune-Albright syndrome, ACTH-independent macronodular hyperplasia, and, rarely, cortisol-producing adenomas. Objective and Design: The whole-genome expression profile (WGEP) of normal (pooled) adrenals, PRKAR1A- (3) and GNAS-mutant (3) was studied. Quantitative RT-PCR and Western blot were used to validate WGEP findings. Results: MAPK and p53 signaling pathways were highly overexpressed in all lesions against normal tissue. GNAS-mutant tissues were significantly enriched for extracellular matrix receptor interaction and focal adhesion pathways when compared with PRKAR1A-mutant (fold enrichment 3.5, P < 0.0001 and 2.1, P < 0.002, respective...

  • mutations and polymorphisms in the gene encoding regulatory subunit type 1 alpha of protein kinase a PRKAR1A an update
    Human Mutation, 2010
    Co-Authors: Anelia Horvath, Kit Man Tsang, Lionel Groussin, Jerome Bertherat, Laure Cazabat, R Libe, Marine Guillaudbataille
    Abstract:

    PRKAR1A encodes the regulatory subunit type 1-alpha (RIalpha) of the cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA). Inactivating PRKAR1A mutations are known to be responsible for the multiple neoplasia and lentiginosis syndrome Carney complex (CNC). To date, at least 117 pathogenic variants in PRKAR1A have been identified (online database: http://PRKAR1A.nichd.nih.gov). The majority are subject to nonsense mediated mRNA decay (NMD), leading to RIalpha haploinsufficiency and, as a result, activated cAMP signaling. Recently, it became apparent that CNC may be caused not only by RIalpha haploinsufficiency, but also by the expression of altered RIalpha protein, as proven by analysis of expressed mutations in the gene, consisting of amino acid substitutions and in-frame genetic alterations. In addition, a new subgroup of mutations that potentially escape NMD and result in CNC through altered (rather than missing) protein has been analyzed-these are frame-shifts in the 3' end of the coding sequence that shift the stop codon downstream of the normal one. The mutation detection rate in CNC patients is recently estimated at above 60%; PRKAR1A mutation-negative CNC patients are characterized by significant phenotypic heterogeneity. In this report, we present a comprehensive analysis of all presently known PRKAR1A sequence variations and discuss their molecular context and clinical phenotype.