Limits...
Coexpression of human somatostatin receptor-2 (SSTR2) and SSTR3 modulates antiproliferative signaling and apoptosis.

War SA, Kumar U - J Mol Signal (2012)

Bottom Line: Although in basal conditions, SSTR2 and SSTR3 colocalize at the plasma membrane and exhibit heterodimerization, the cell surface distribution of both receptors decreased upon agonist activation and was accompanied by a parallel increase in intracellular colocalization.Receptors activation by SST and specific agonists significantly decreased cAMP levels in cotransfected cells in comparison to control.The receptor activation in cotransfected cells exhibits Gi-dependent inhibition of cell proliferation attributed to increased PARP-1 expression and TUNEL staining, whereas induction of p21 and p27Kip1 suggests a cytostatic effect.

View Article: PubMed Central - HTML - PubMed

Affiliation: Faculty of Pharmaceutical Sciences, The University of British Columbia, Vancouver, BC, V6T 1Z3, Canada. ujkumar@mail.ubc.ca.

ABSTRACT

Background: Somatostatin (SST) via five Gi coupled receptors namely SSTR1-5 is known to inhibit cell proliferation by cytostatic and cytotoxic mechanisms. Heterodimerization plays a crucial role in modulating the signal transduction pathways of SSTR subtypes. In the present study, we investigated human SSTR2/SSTR3 heterodimerization, internalization, MAPK signaling, cell proliferation and apoptosis in HEK-293 cells in response to SST and specific agonists for SSTR2 and SSTR3.

Results: Although in basal conditions, SSTR2 and SSTR3 colocalize at the plasma membrane and exhibit heterodimerization, the cell surface distribution of both receptors decreased upon agonist activation and was accompanied by a parallel increase in intracellular colocalization. Receptors activation by SST and specific agonists significantly decreased cAMP levels in cotransfected cells in comparison to control. Agonist-mediated modulation of pERK1/2 was time and concentration-dependent, and pronounced in serum-deprived conditions. pERK1/2 was inhibited in response to SST; conversely receptor-specific agonist treatment caused inhibition at lower concentration and activation at higher concentration. Strikingly, ERK1/2 phosphorylation was sustained upon prolonged treatment with SST but not with receptor-specific agonists. On the other hand, SST and receptor-specific agonists modulated p38 phosphorylation time-dependently. The receptor activation in cotransfected cells exhibits Gi-dependent inhibition of cell proliferation attributed to increased PARP-1 expression and TUNEL staining, whereas induction of p21 and p27Kip1 suggests a cytostatic effect.

Conclusion: Our study provides new insights in SSTR2/SSTR3 mediated signaling which might help in better understanding of the molecular interactions involving SSTRs in tumor biology.

No MeSH data available.


Related in: MedlinePlus

Concentration and time-dependent modulation of ERK1/2 phosphorylation by SSTR2/SSTR3. Cells stably cotransfected with SSTR2 and SSTR3 were treated with SST (1, 50 nM and 1 μM), L-779976 (10 and 100 nM) and L-796778 (25 and 50 nM) for 30 min (top panel) or 24 h (bottom panel) with or without FBS, and processed for total and pERK1/2 using Western blot analysis. Note a distinct pattern of ERK1/2 phosphorylation in response to SST and receptor-specific agonists in a concentration and time-dependent manner. Histograms represent the densitometric analysis and the results are presented as the ratio of phospho- and total-ERK1/2 expressed as fold over control. Mean ± S.D is representative of three independent experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3403965&req=5

Figure 4: Concentration and time-dependent modulation of ERK1/2 phosphorylation by SSTR2/SSTR3. Cells stably cotransfected with SSTR2 and SSTR3 were treated with SST (1, 50 nM and 1 μM), L-779976 (10 and 100 nM) and L-796778 (25 and 50 nM) for 30 min (top panel) or 24 h (bottom panel) with or without FBS, and processed for total and pERK1/2 using Western blot analysis. Note a distinct pattern of ERK1/2 phosphorylation in response to SST and receptor-specific agonists in a concentration and time-dependent manner. Histograms represent the densitometric analysis and the results are presented as the ratio of phospho- and total-ERK1/2 expressed as fold over control. Mean ± S.D is representative of three independent experiments.

Mentions: Our recent studies suggested agonist and time-dependent modulation of pERK1/2 in cells expressing SSTR2, SSTR3 or SSTR2/SSTR5 [3,9]. Using time-course experiments (30 min and 24 h), we examined the effect of heterodimerization on ERK1/2 phosphorylation in cotransfected cells with or without FBS. As depicted in Figure 4, the levels of pERK1/2 were significantly increased at lower and higher concentrations of SST for 30 min; whereas in response to receptor-specific agonists, ERK1/2 phosphorylation decreased at lower concentration and increased at higher concentration when compared to control (*, p < 0.05; **, p < 0.01). In the absence of FBS, pERK1/2 was significantly decreased by SST and receptor-specific agonists in comparison to control (*, p < 0.05). SST treatment for 24 h increased pERK1/2 when compared to non-treated cells (*, p < 0.05), whereas no significant changes were observed with receptor-specific agonists alone suggesting that co-activation of both receptors is a prerequisite for sustained ERK1/2 signaling. However in FBS-deficient conditions, SST and receptor-specific agonists significantly enhanced pERK1/2 in comparison to control (*, p < 0.05).


Coexpression of human somatostatin receptor-2 (SSTR2) and SSTR3 modulates antiproliferative signaling and apoptosis.

War SA, Kumar U - J Mol Signal (2012)

Concentration and time-dependent modulation of ERK1/2 phosphorylation by SSTR2/SSTR3. Cells stably cotransfected with SSTR2 and SSTR3 were treated with SST (1, 50 nM and 1 μM), L-779976 (10 and 100 nM) and L-796778 (25 and 50 nM) for 30 min (top panel) or 24 h (bottom panel) with or without FBS, and processed for total and pERK1/2 using Western blot analysis. Note a distinct pattern of ERK1/2 phosphorylation in response to SST and receptor-specific agonists in a concentration and time-dependent manner. Histograms represent the densitometric analysis and the results are presented as the ratio of phospho- and total-ERK1/2 expressed as fold over control. Mean ± S.D is representative of three independent experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3403965&req=5

Figure 4: Concentration and time-dependent modulation of ERK1/2 phosphorylation by SSTR2/SSTR3. Cells stably cotransfected with SSTR2 and SSTR3 were treated with SST (1, 50 nM and 1 μM), L-779976 (10 and 100 nM) and L-796778 (25 and 50 nM) for 30 min (top panel) or 24 h (bottom panel) with or without FBS, and processed for total and pERK1/2 using Western blot analysis. Note a distinct pattern of ERK1/2 phosphorylation in response to SST and receptor-specific agonists in a concentration and time-dependent manner. Histograms represent the densitometric analysis and the results are presented as the ratio of phospho- and total-ERK1/2 expressed as fold over control. Mean ± S.D is representative of three independent experiments.
Mentions: Our recent studies suggested agonist and time-dependent modulation of pERK1/2 in cells expressing SSTR2, SSTR3 or SSTR2/SSTR5 [3,9]. Using time-course experiments (30 min and 24 h), we examined the effect of heterodimerization on ERK1/2 phosphorylation in cotransfected cells with or without FBS. As depicted in Figure 4, the levels of pERK1/2 were significantly increased at lower and higher concentrations of SST for 30 min; whereas in response to receptor-specific agonists, ERK1/2 phosphorylation decreased at lower concentration and increased at higher concentration when compared to control (*, p < 0.05; **, p < 0.01). In the absence of FBS, pERK1/2 was significantly decreased by SST and receptor-specific agonists in comparison to control (*, p < 0.05). SST treatment for 24 h increased pERK1/2 when compared to non-treated cells (*, p < 0.05), whereas no significant changes were observed with receptor-specific agonists alone suggesting that co-activation of both receptors is a prerequisite for sustained ERK1/2 signaling. However in FBS-deficient conditions, SST and receptor-specific agonists significantly enhanced pERK1/2 in comparison to control (*, p < 0.05).

Bottom Line: Although in basal conditions, SSTR2 and SSTR3 colocalize at the plasma membrane and exhibit heterodimerization, the cell surface distribution of both receptors decreased upon agonist activation and was accompanied by a parallel increase in intracellular colocalization.Receptors activation by SST and specific agonists significantly decreased cAMP levels in cotransfected cells in comparison to control.The receptor activation in cotransfected cells exhibits Gi-dependent inhibition of cell proliferation attributed to increased PARP-1 expression and TUNEL staining, whereas induction of p21 and p27Kip1 suggests a cytostatic effect.

View Article: PubMed Central - HTML - PubMed

Affiliation: Faculty of Pharmaceutical Sciences, The University of British Columbia, Vancouver, BC, V6T 1Z3, Canada. ujkumar@mail.ubc.ca.

ABSTRACT

Background: Somatostatin (SST) via five Gi coupled receptors namely SSTR1-5 is known to inhibit cell proliferation by cytostatic and cytotoxic mechanisms. Heterodimerization plays a crucial role in modulating the signal transduction pathways of SSTR subtypes. In the present study, we investigated human SSTR2/SSTR3 heterodimerization, internalization, MAPK signaling, cell proliferation and apoptosis in HEK-293 cells in response to SST and specific agonists for SSTR2 and SSTR3.

Results: Although in basal conditions, SSTR2 and SSTR3 colocalize at the plasma membrane and exhibit heterodimerization, the cell surface distribution of both receptors decreased upon agonist activation and was accompanied by a parallel increase in intracellular colocalization. Receptors activation by SST and specific agonists significantly decreased cAMP levels in cotransfected cells in comparison to control. Agonist-mediated modulation of pERK1/2 was time and concentration-dependent, and pronounced in serum-deprived conditions. pERK1/2 was inhibited in response to SST; conversely receptor-specific agonist treatment caused inhibition at lower concentration and activation at higher concentration. Strikingly, ERK1/2 phosphorylation was sustained upon prolonged treatment with SST but not with receptor-specific agonists. On the other hand, SST and receptor-specific agonists modulated p38 phosphorylation time-dependently. The receptor activation in cotransfected cells exhibits Gi-dependent inhibition of cell proliferation attributed to increased PARP-1 expression and TUNEL staining, whereas induction of p21 and p27Kip1 suggests a cytostatic effect.

Conclusion: Our study provides new insights in SSTR2/SSTR3 mediated signaling which might help in better understanding of the molecular interactions involving SSTRs in tumor biology.

No MeSH data available.


Related in: MedlinePlus