The Experts below are selected from a list of 296640 Experts worldwide ranked by ideXlab platform
Gary L Firestone - One of the best experts on this subject based on the ideXlab platform.
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indole 3 carbinol downregulation of telomerase gene expression requires the inhibition of estrogen receptor alpha and sp1 transcription factor interactions within the htert promoter and mediates the g1 cell cycle arrest of human breast cancer cells
2011Co-Authors: Crystal N Marconett, Shyam N Sundar, Kalvin Q Tran, Kelly Mahuron, Min Tseng, Leonard F Bjeldanes, Gary L FirestoneAbstract:Indole-3-carbinol (I3C), a naturally occurring hydrolysis product of glucobrassicin from cruciferous vegetables such as broccoli, cabbage and Brussels sprouts, is an anticancer phytochemical that triggers complementary sets of antiproliferative pathways to induce a cell cycle arrest of estrogen-responsive MCF7 breast cancer cells. I3C strongly downregulated transcript expression of the catalytic subunit of the human telomerase (hTERT) gene, which correlated with the dose-dependent indole-mediated G1 cell cycle arrest without altering the transcript levels of the RNA template (hTR) for telomerase elongation. Exogenous expression of hTERT driven by a constitutive promoter prevented the I3C-induced cell cycle arrest and rescued the I3C inhibition of telomerase enzymatic activity and activation of cellular senescence. Time course studies showed that I3C downregulated expression of estrogen receptor-alpha (ERα) and Cyclin-Dependent Kinase-6 transcripts levels (which is regulated through the Sp1 transcription factor) prior to the downregulation of hTERT suggesting a mechanistic link. Chromatin immunoprecipitation assays demonstrated that I3C disrupted endogenous interactions of both ERα and Sp1 with an estrogen response element–Sp1 composite element within the hTERT promoter. I3C inhibited 17β-estradiol stimulated hTERT expression and stimulated the production of threonine-phosphorylated Sp1, which inhibits Sp1–DNA interactions. Exogenous expression of both ERα and Sp1, but not either alone, in MCF7 cells blocked the I3C-mediated downregulation of hTERT expression. These results demonstrate that I3C disrupts the combined ERα- and Sp1-driven transcription of hTERT gene expression, which plays a significant role in the I3C-induced cell cycle arrest of human breast cancer cells.
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indole 3 carbinol and 3 3 diindolylmethane antiproliferative signaling pathways control cell cycle gene transcription in human breast cancer cells by regulating promoter sp1 transcription factor interactions
2003Co-Authors: Gary L Firestone, Leonard F BjeldanesAbstract:Indole-3-carbinol (I3C), a compound that occurs naturally in Brassica vegetables such as cabbage and broccoli, can induce a G1 cell-cycle arrest of human MCF-7 breast cancer cells that is accompanied by the selective inhibition of Cyclin-Dependent Kinase 6 (Cdk6) expression and stimulation of p21(Waf1/Cip1) gene expression. Construction and transfection of a series of promoter-reporter plasmids demonstrate that the indole-regulated changes in Cdk6 and p21(Waf1/Cip1) levels are due to specific effects on their corresponding promoters. Mutagenic analysis reveals that I3C signaling targets a composite transcriptional element in the Cdk6 promoter that requires both Sp1 and Ets transcription factors for transactivation function. Analysis of protein-DNA complexes formed with nuclear proteins isolated from I3C-treated and -untreated cells demonstrates that the Sp1 DNA element in the Cdk6 promoter interacts with an I3C-inhibited protein-protein complex that contains the Sp1 transcription factor. In indole-treated cells, a fraction of [(3)H]I3C was converted into its natural diindole product (3)H-labeled 3-3'-diindolylmethane ([(3)H]DIM), which accumulates in the nucleus; this suggests that DIM may have a role in the transcriptional activities of I3C. Mutagenic analysis of the p21(Waf1/Cip1) promoter reveals that in transfected breast cancer cells, DIM (as well as I3C) stimulates p21(Waf1/Cip1) transcription through an indole-responsive region of the promoter that contains multiple Sp1 consensus sequences. Furthermore, DIM treatment regulates the presence of a nuclear Sp1 DNA-binding activity. Our results demonstrate that both the Cdk6 and p21(Waf1/Cip1) promoters are newly defined downstream targets of the indole-signaling pathway, and that the observed transcriptional effects are due to a combination of the cellular activities of I3C and DIM.
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indole 3 carbinol inhibits cdk6 expression in human mcf 7 breast cancer cells by disrupting sp1 transcription factor interactions with a composite element in the cdk6 gene promoter
2001Co-Authors: Erin J. Cram, Leonard F Bjeldanes, Gary L FirestoneAbstract:Abstract Indole-3-carbinol (I3C), a compound naturally occurring in Brassica vegetables, can induce a G1 cell cycle arrest of human MCF-7 breast cancer cells that is accompanied by the selective inhibition of Cyclin-Dependent Kinase 6 (CDK6) expression. Reverse transcriptase-polymerase chain reaction analysis of CDK6 mRNA decay rates revealed that I3C had no effect on CDK6 transcript stability. We report the first identification and functional characterization of theCDK6 promoter in order to determine whether I3C inhibits CDK6 transcription. In MCF-7 cells stably transfected withCDK6 promoter-linked luciferase reporter plasmids, I3C inhibited CDK6 promoter activity in an I3C-specific response that was not a consequence of the growth-arrested state of the cells. Deletion analysis revealed a 167-base pair I3C-responsive region of the CDK6 promoter between −805 and −638. Site-specific mutations within this region revealed that both Sp1 and Ets-like sites, which are spaced 5 base pairs apart, were necessary for I3C responsiveness in the context of the CDK6 promoter. Electrophoretic mobility shift analysis of protein-DNA complexes formed with nuclear proteins isolated from I3C-treated and -untreated cells, in combination with supershift assays using Sp1 antibodies, demonstrated that the Sp1-binding site in the CDK6 promoter forms a specific I3C-responsive DNA-protein complex that contains the Sp1 transcription factor. Taken together, our results suggest that I3C down-regulates CDK6 transcription by targeting Sp1 at a composite DNA site in the CDK6 promoter.
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indole 3 carbinol inhibits the expression of cyclin dependent Kinase 6 and induces a g1 cell cycle arrest of human breast cancer cells independent of estrogen receptor signaling
1998Co-Authors: Carolyn M Cover, Leonard F Bjeldanes, Jean S Hsieh, Susan H Tran, Gunnell Hallden, Gloria S Kim, Gary L FirestoneAbstract:Indole-3-carbinol (I3C), a naturally occurring component of Brassica vegetables such as cabbage, broccoli, and Brussels sprouts, has been shown to reduce the incidence of spontaneous and carcinogen-induced mammary tumors. Treatment of cultured human MCF7 breast cancer cells with I3C reversibly suppresses the incorporation of [3H]thymidine without affecting cell viability or estrogen receptor (ER) responsiveness. Flow cytometry of propidium iodide-stained cells revealed that I3C induces a G1 cell cycle arrest. Concurrent with the I3C-induced growth inhibition, Northern blot and Western blot analyses demonstrated that I3C selectively abolished the expression of Cyclin-Dependent Kinase 6 (CDK6) in a dose- and time-dependent manner. Furthermore, I3C inhibited the endogenous retinoblastoma protein phosphorylation and CDK6 phosphorylation of retinoblastoma in vitro to the same extent. After the MCF7 cells reached their maximal growth arrest, the levels of the p21 and p27 CDK inhibitors increased by 50%. The antiestrogen tamoxifen also suppressed MCF7 cell DNA synthesis but had no effect on CDK6 expression, while a combination of I3C and tamoxifen inhibited MCF7 cell growth more stringently than either agent alone. The I3C-mediated cell cycle arrest and repression of CDK6 production were also observed in estrogen receptor-deficient MDA-MB-231 human breast cancer cells, which demonstrates that this indole can suppress the growth of mammary tumor cells independent of estrogen receptor signaling. Thus, our observations have uncovered a previously undefined antiproliferative pathway for I3C that implicates CDK6 as a target for cell cycle control in human breast cancer cells. Moreover, our results establish for the first time that CDK6 gene expression can be inhibited in response to an extracellular antiproliferative signal.
Caiqiao Zhang - One of the best experts on this subject based on the ideXlab platform.
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retinoic acid promotes proliferation of chicken primordial germ cells via activation of pi3k akt mediated nf κb signalling cascade
2012Co-Authors: Minli Yu, Chutian Ge, Yuling Mi, Weidong Zeng, Caiqiao ZhangAbstract:: As embryonic progenitors for the gametes, PGCs (primordial germ cells) proliferate and develop under strict regulation of numerous intrinsic and external factors. As the most active natural metabolite of vitamin A, all-trans RA (retinoic acid) plays pivotal roles in regulating development of various cells. The proliferating action of RA on PGCs was investigated along with the intracellular PI3K (phosphoinositide 3-Kinase)/Akt (protein Kinase B; also known as Akt)-mediated NF-κB (nuclear factor κB) signalling cascade. The results show that RA significantly promoted PGC proliferation in a dose- and time-dependent manner, confirmed by BrdU (bromodeoxyuridine) incorporation and cell cycle analysis. However, this promoting effect was attenuated by sequential inhibitors of LY294002 for PI3K, KP372-1 for Akt and SN50 for NF-κB respectively. Western blot analysis showed increased Akt phosphorylation (Ser473) of PGCs after stimulation with RA, but this was abolished by LY294002 or KP372-1. Treatment with RA increased expression of NF-κB and decreased IκBα (inhibitory κBα) expression, which were inhibited by SN50. Blockade of PI3K or Akt activity inhibited NF-κB translocation from the cytoplasm to the nucleus. Finally, mRNA expression of cell cycle regulating genes [cyclin D1 and E, CDK6 (Cyclin-Dependent Kinase 6) and CDK2] was up-regulated in the RA-treated cells. This stimulation was also markedly retarded by combined treatment with LY294002, KP372-1 and SN50. These results suggest that RA activates the PI3K/Akt and NF-κB signalling cascade to promote proliferation of the cultured chicken PGCs.
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retinoic acid promotes proliferation of chicken primordial germ cells via activation of pi3k akt mediated nf κb signalling cascade
2012Co-Authors: Minli Yu, Chutian Ge, Yuling Mi, Weidong Zeng, Caiqiao ZhangAbstract:: As embryonic progenitors for the gametes, PGCs (primordial germ cells) proliferate and develop under strict regulation of numerous intrinsic and external factors. As the most active natural metabolite of vitamin A, all-trans RA (retinoic acid) plays pivotal roles in regulating development of various cells. The proliferating action of RA on PGCs was investigated along with the intracellular PI3K (phosphoinositide 3-Kinase)/Akt (protein Kinase B; also known as Akt)-mediated NF-κB (nuclear factor κB) signalling cascade. The results show that RA significantly promoted PGC proliferation in a dose- and time-dependent manner, confirmed by BrdU (bromodeoxyuridine) incorporation and cell cycle analysis. However, this promoting effect was attenuated by sequential inhibitors of LY294002 for PI3K, KP372-1 for Akt and SN50 for NF-κB respectively. Western blot analysis showed increased Akt phosphorylation (Ser473) of PGCs after stimulation with RA, but this was abolished by LY294002 or KP372-1. Treatment with RA increased expression of NF-κB and decreased IκBα (inhibitory κBα) expression, which were inhibited by SN50. Blockade of PI3K or Akt activity inhibited NF-κB translocation from the cytoplasm to the nucleus. Finally, mRNA expression of cell cycle regulating genes [cyclin D1 and E, CDK6 (Cyclin-Dependent Kinase 6) and CDK2] was up-regulated in the RA-treated cells. This stimulation was also markedly retarded by combined treatment with LY294002, KP372-1 and SN50. These results suggest that RA activates the PI3K/Akt and NF-κB signalling cascade to promote proliferation of the cultured chicken PGCs.
Leonard F Bjeldanes - One of the best experts on this subject based on the ideXlab platform.
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indole 3 carbinol downregulation of telomerase gene expression requires the inhibition of estrogen receptor alpha and sp1 transcription factor interactions within the htert promoter and mediates the g1 cell cycle arrest of human breast cancer cells
2011Co-Authors: Crystal N Marconett, Shyam N Sundar, Kalvin Q Tran, Kelly Mahuron, Min Tseng, Leonard F Bjeldanes, Gary L FirestoneAbstract:Indole-3-carbinol (I3C), a naturally occurring hydrolysis product of glucobrassicin from cruciferous vegetables such as broccoli, cabbage and Brussels sprouts, is an anticancer phytochemical that triggers complementary sets of antiproliferative pathways to induce a cell cycle arrest of estrogen-responsive MCF7 breast cancer cells. I3C strongly downregulated transcript expression of the catalytic subunit of the human telomerase (hTERT) gene, which correlated with the dose-dependent indole-mediated G1 cell cycle arrest without altering the transcript levels of the RNA template (hTR) for telomerase elongation. Exogenous expression of hTERT driven by a constitutive promoter prevented the I3C-induced cell cycle arrest and rescued the I3C inhibition of telomerase enzymatic activity and activation of cellular senescence. Time course studies showed that I3C downregulated expression of estrogen receptor-alpha (ERα) and Cyclin-Dependent Kinase-6 transcripts levels (which is regulated through the Sp1 transcription factor) prior to the downregulation of hTERT suggesting a mechanistic link. Chromatin immunoprecipitation assays demonstrated that I3C disrupted endogenous interactions of both ERα and Sp1 with an estrogen response element–Sp1 composite element within the hTERT promoter. I3C inhibited 17β-estradiol stimulated hTERT expression and stimulated the production of threonine-phosphorylated Sp1, which inhibits Sp1–DNA interactions. Exogenous expression of both ERα and Sp1, but not either alone, in MCF7 cells blocked the I3C-mediated downregulation of hTERT expression. These results demonstrate that I3C disrupts the combined ERα- and Sp1-driven transcription of hTERT gene expression, which plays a significant role in the I3C-induced cell cycle arrest of human breast cancer cells.
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indole 3 carbinol and 3 3 diindolylmethane antiproliferative signaling pathways control cell cycle gene transcription in human breast cancer cells by regulating promoter sp1 transcription factor interactions
2003Co-Authors: Gary L Firestone, Leonard F BjeldanesAbstract:Indole-3-carbinol (I3C), a compound that occurs naturally in Brassica vegetables such as cabbage and broccoli, can induce a G1 cell-cycle arrest of human MCF-7 breast cancer cells that is accompanied by the selective inhibition of Cyclin-Dependent Kinase 6 (Cdk6) expression and stimulation of p21(Waf1/Cip1) gene expression. Construction and transfection of a series of promoter-reporter plasmids demonstrate that the indole-regulated changes in Cdk6 and p21(Waf1/Cip1) levels are due to specific effects on their corresponding promoters. Mutagenic analysis reveals that I3C signaling targets a composite transcriptional element in the Cdk6 promoter that requires both Sp1 and Ets transcription factors for transactivation function. Analysis of protein-DNA complexes formed with nuclear proteins isolated from I3C-treated and -untreated cells demonstrates that the Sp1 DNA element in the Cdk6 promoter interacts with an I3C-inhibited protein-protein complex that contains the Sp1 transcription factor. In indole-treated cells, a fraction of [(3)H]I3C was converted into its natural diindole product (3)H-labeled 3-3'-diindolylmethane ([(3)H]DIM), which accumulates in the nucleus; this suggests that DIM may have a role in the transcriptional activities of I3C. Mutagenic analysis of the p21(Waf1/Cip1) promoter reveals that in transfected breast cancer cells, DIM (as well as I3C) stimulates p21(Waf1/Cip1) transcription through an indole-responsive region of the promoter that contains multiple Sp1 consensus sequences. Furthermore, DIM treatment regulates the presence of a nuclear Sp1 DNA-binding activity. Our results demonstrate that both the Cdk6 and p21(Waf1/Cip1) promoters are newly defined downstream targets of the indole-signaling pathway, and that the observed transcriptional effects are due to a combination of the cellular activities of I3C and DIM.
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indole 3 carbinol inhibits cdk6 expression in human mcf 7 breast cancer cells by disrupting sp1 transcription factor interactions with a composite element in the cdk6 gene promoter
2001Co-Authors: Erin J. Cram, Leonard F Bjeldanes, Gary L FirestoneAbstract:Abstract Indole-3-carbinol (I3C), a compound naturally occurring in Brassica vegetables, can induce a G1 cell cycle arrest of human MCF-7 breast cancer cells that is accompanied by the selective inhibition of Cyclin-Dependent Kinase 6 (CDK6) expression. Reverse transcriptase-polymerase chain reaction analysis of CDK6 mRNA decay rates revealed that I3C had no effect on CDK6 transcript stability. We report the first identification and functional characterization of theCDK6 promoter in order to determine whether I3C inhibits CDK6 transcription. In MCF-7 cells stably transfected withCDK6 promoter-linked luciferase reporter plasmids, I3C inhibited CDK6 promoter activity in an I3C-specific response that was not a consequence of the growth-arrested state of the cells. Deletion analysis revealed a 167-base pair I3C-responsive region of the CDK6 promoter between −805 and −638. Site-specific mutations within this region revealed that both Sp1 and Ets-like sites, which are spaced 5 base pairs apart, were necessary for I3C responsiveness in the context of the CDK6 promoter. Electrophoretic mobility shift analysis of protein-DNA complexes formed with nuclear proteins isolated from I3C-treated and -untreated cells, in combination with supershift assays using Sp1 antibodies, demonstrated that the Sp1-binding site in the CDK6 promoter forms a specific I3C-responsive DNA-protein complex that contains the Sp1 transcription factor. Taken together, our results suggest that I3C down-regulates CDK6 transcription by targeting Sp1 at a composite DNA site in the CDK6 promoter.
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indole 3 carbinol inhibits the expression of cyclin dependent Kinase 6 and induces a g1 cell cycle arrest of human breast cancer cells independent of estrogen receptor signaling
1998Co-Authors: Carolyn M Cover, Leonard F Bjeldanes, Jean S Hsieh, Susan H Tran, Gunnell Hallden, Gloria S Kim, Gary L FirestoneAbstract:Indole-3-carbinol (I3C), a naturally occurring component of Brassica vegetables such as cabbage, broccoli, and Brussels sprouts, has been shown to reduce the incidence of spontaneous and carcinogen-induced mammary tumors. Treatment of cultured human MCF7 breast cancer cells with I3C reversibly suppresses the incorporation of [3H]thymidine without affecting cell viability or estrogen receptor (ER) responsiveness. Flow cytometry of propidium iodide-stained cells revealed that I3C induces a G1 cell cycle arrest. Concurrent with the I3C-induced growth inhibition, Northern blot and Western blot analyses demonstrated that I3C selectively abolished the expression of Cyclin-Dependent Kinase 6 (CDK6) in a dose- and time-dependent manner. Furthermore, I3C inhibited the endogenous retinoblastoma protein phosphorylation and CDK6 phosphorylation of retinoblastoma in vitro to the same extent. After the MCF7 cells reached their maximal growth arrest, the levels of the p21 and p27 CDK inhibitors increased by 50%. The antiestrogen tamoxifen also suppressed MCF7 cell DNA synthesis but had no effect on CDK6 expression, while a combination of I3C and tamoxifen inhibited MCF7 cell growth more stringently than either agent alone. The I3C-mediated cell cycle arrest and repression of CDK6 production were also observed in estrogen receptor-deficient MDA-MB-231 human breast cancer cells, which demonstrates that this indole can suppress the growth of mammary tumor cells independent of estrogen receptor signaling. Thus, our observations have uncovered a previously undefined antiproliferative pathway for I3C that implicates CDK6 as a target for cell cycle control in human breast cancer cells. Moreover, our results establish for the first time that CDK6 gene expression can be inhibited in response to an extracellular antiproliferative signal.
Minli Yu - One of the best experts on this subject based on the ideXlab platform.
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retinoic acid promotes proliferation of chicken primordial germ cells via activation of pi3k akt mediated nf κb signalling cascade
2012Co-Authors: Minli Yu, Chutian Ge, Yuling Mi, Weidong Zeng, Caiqiao ZhangAbstract:: As embryonic progenitors for the gametes, PGCs (primordial germ cells) proliferate and develop under strict regulation of numerous intrinsic and external factors. As the most active natural metabolite of vitamin A, all-trans RA (retinoic acid) plays pivotal roles in regulating development of various cells. The proliferating action of RA on PGCs was investigated along with the intracellular PI3K (phosphoinositide 3-Kinase)/Akt (protein Kinase B; also known as Akt)-mediated NF-κB (nuclear factor κB) signalling cascade. The results show that RA significantly promoted PGC proliferation in a dose- and time-dependent manner, confirmed by BrdU (bromodeoxyuridine) incorporation and cell cycle analysis. However, this promoting effect was attenuated by sequential inhibitors of LY294002 for PI3K, KP372-1 for Akt and SN50 for NF-κB respectively. Western blot analysis showed increased Akt phosphorylation (Ser473) of PGCs after stimulation with RA, but this was abolished by LY294002 or KP372-1. Treatment with RA increased expression of NF-κB and decreased IκBα (inhibitory κBα) expression, which were inhibited by SN50. Blockade of PI3K or Akt activity inhibited NF-κB translocation from the cytoplasm to the nucleus. Finally, mRNA expression of cell cycle regulating genes [cyclin D1 and E, CDK6 (Cyclin-Dependent Kinase 6) and CDK2] was up-regulated in the RA-treated cells. This stimulation was also markedly retarded by combined treatment with LY294002, KP372-1 and SN50. These results suggest that RA activates the PI3K/Akt and NF-κB signalling cascade to promote proliferation of the cultured chicken PGCs.
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retinoic acid promotes proliferation of chicken primordial germ cells via activation of pi3k akt mediated nf κb signalling cascade
2012Co-Authors: Minli Yu, Chutian Ge, Yuling Mi, Weidong Zeng, Caiqiao ZhangAbstract:: As embryonic progenitors for the gametes, PGCs (primordial germ cells) proliferate and develop under strict regulation of numerous intrinsic and external factors. As the most active natural metabolite of vitamin A, all-trans RA (retinoic acid) plays pivotal roles in regulating development of various cells. The proliferating action of RA on PGCs was investigated along with the intracellular PI3K (phosphoinositide 3-Kinase)/Akt (protein Kinase B; also known as Akt)-mediated NF-κB (nuclear factor κB) signalling cascade. The results show that RA significantly promoted PGC proliferation in a dose- and time-dependent manner, confirmed by BrdU (bromodeoxyuridine) incorporation and cell cycle analysis. However, this promoting effect was attenuated by sequential inhibitors of LY294002 for PI3K, KP372-1 for Akt and SN50 for NF-κB respectively. Western blot analysis showed increased Akt phosphorylation (Ser473) of PGCs after stimulation with RA, but this was abolished by LY294002 or KP372-1. Treatment with RA increased expression of NF-κB and decreased IκBα (inhibitory κBα) expression, which were inhibited by SN50. Blockade of PI3K or Akt activity inhibited NF-κB translocation from the cytoplasm to the nucleus. Finally, mRNA expression of cell cycle regulating genes [cyclin D1 and E, CDK6 (Cyclin-Dependent Kinase 6) and CDK2] was up-regulated in the RA-treated cells. This stimulation was also markedly retarded by combined treatment with LY294002, KP372-1 and SN50. These results suggest that RA activates the PI3K/Akt and NF-κB signalling cascade to promote proliferation of the cultured chicken PGCs.
Yuling Mi - One of the best experts on this subject based on the ideXlab platform.
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retinoic acid promotes proliferation of chicken primordial germ cells via activation of pi3k akt mediated nf κb signalling cascade
2012Co-Authors: Minli Yu, Chutian Ge, Yuling Mi, Weidong Zeng, Caiqiao ZhangAbstract:: As embryonic progenitors for the gametes, PGCs (primordial germ cells) proliferate and develop under strict regulation of numerous intrinsic and external factors. As the most active natural metabolite of vitamin A, all-trans RA (retinoic acid) plays pivotal roles in regulating development of various cells. The proliferating action of RA on PGCs was investigated along with the intracellular PI3K (phosphoinositide 3-Kinase)/Akt (protein Kinase B; also known as Akt)-mediated NF-κB (nuclear factor κB) signalling cascade. The results show that RA significantly promoted PGC proliferation in a dose- and time-dependent manner, confirmed by BrdU (bromodeoxyuridine) incorporation and cell cycle analysis. However, this promoting effect was attenuated by sequential inhibitors of LY294002 for PI3K, KP372-1 for Akt and SN50 for NF-κB respectively. Western blot analysis showed increased Akt phosphorylation (Ser473) of PGCs after stimulation with RA, but this was abolished by LY294002 or KP372-1. Treatment with RA increased expression of NF-κB and decreased IκBα (inhibitory κBα) expression, which were inhibited by SN50. Blockade of PI3K or Akt activity inhibited NF-κB translocation from the cytoplasm to the nucleus. Finally, mRNA expression of cell cycle regulating genes [cyclin D1 and E, CDK6 (Cyclin-Dependent Kinase 6) and CDK2] was up-regulated in the RA-treated cells. This stimulation was also markedly retarded by combined treatment with LY294002, KP372-1 and SN50. These results suggest that RA activates the PI3K/Akt and NF-κB signalling cascade to promote proliferation of the cultured chicken PGCs.
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retinoic acid promotes proliferation of chicken primordial germ cells via activation of pi3k akt mediated nf κb signalling cascade
2012Co-Authors: Minli Yu, Chutian Ge, Yuling Mi, Weidong Zeng, Caiqiao ZhangAbstract:: As embryonic progenitors for the gametes, PGCs (primordial germ cells) proliferate and develop under strict regulation of numerous intrinsic and external factors. As the most active natural metabolite of vitamin A, all-trans RA (retinoic acid) plays pivotal roles in regulating development of various cells. The proliferating action of RA on PGCs was investigated along with the intracellular PI3K (phosphoinositide 3-Kinase)/Akt (protein Kinase B; also known as Akt)-mediated NF-κB (nuclear factor κB) signalling cascade. The results show that RA significantly promoted PGC proliferation in a dose- and time-dependent manner, confirmed by BrdU (bromodeoxyuridine) incorporation and cell cycle analysis. However, this promoting effect was attenuated by sequential inhibitors of LY294002 for PI3K, KP372-1 for Akt and SN50 for NF-κB respectively. Western blot analysis showed increased Akt phosphorylation (Ser473) of PGCs after stimulation with RA, but this was abolished by LY294002 or KP372-1. Treatment with RA increased expression of NF-κB and decreased IκBα (inhibitory κBα) expression, which were inhibited by SN50. Blockade of PI3K or Akt activity inhibited NF-κB translocation from the cytoplasm to the nucleus. Finally, mRNA expression of cell cycle regulating genes [cyclin D1 and E, CDK6 (Cyclin-Dependent Kinase 6) and CDK2] was up-regulated in the RA-treated cells. This stimulation was also markedly retarded by combined treatment with LY294002, KP372-1 and SN50. These results suggest that RA activates the PI3K/Akt and NF-κB signalling cascade to promote proliferation of the cultured chicken PGCs.
