Smad7 is a transforming growth factor-beta-inducible mediator of apoptosis in granulosa cells.
OBJECTIVE
To determine the functional role of Smad7 in granulosa cells.


METHODS
Granulosa cell culture and molecular biological techniques were used to investigate regulation and function of Smad7.


METHODS
Research laboratory.


METHODS
C57bl/j hybrid mouse.


METHODS
Primary mouse granulosa cells were isolated and grown in culture for all messenger RNA expression experiments. Smad7 promoter constructs were evaluated with a luciferase reporter system in SIGC cells to determine sites activating Smad7 expression.


METHODS
Overexpression (Smad7 complementary DNA) and downregulation (Smad7 small interfering RNA) of Smad7 in primary mouse granulosa cells were used to evaluate the functional role of Smad7 in granulosa cells.


RESULTS
Smad7 expression was upregulated by treatment with transforming growth factor-β (TGF-β) but not activin or activation of the cyclic adenosine monophosphate pathway. The promoter of Smad7 was activated by TGF-β. Truncation of the promoter or mutation of the Smad response element at -141 eliminated TGF-β activation of the promoter. Smad3 was not specifically required for TGF-β-stimulated expression of Smad7, though activation of the TGFBR1 receptor was. When Smad7 was overexpressed in granulosa cells, apoptosis was markedly increased. When Smad7 expression was reduced with small interfering RNA, then the TGF-β-induced apoptosis was blocked.


CONCLUSIONS
Smad7 mediates apoptosis induced by TGF-β in mouse granulosa cells, suggesting that dysregulation of Smad7 could impair folliculogenesis.
Department of Obstetrics and Gynecology and Institute for Women's Health, Virginia Commonwealth University, Richmond, Virginia 23298-0034, USA.
Marisol
M
Quezada
Jikui
J
Wang
Valerie
V
Hoang
Elizabeth A
EA
McGee
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D005260:Female; D006107:Granulosa Cells / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D008156:Luciferases / Q000235:genetics; D051379:Mice; D008810:Mice, Inbred C57BL; D009866:Oogenesis / Q000502:physiology*; D061251:Primary Cell Culture; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D051906:Smad7 Protein / Q000235:genetics*; D016212:Transforming Growth Factor beta / Q000494:pharmacology*
D012333:RNA, Messenger; D034741:RNA, Small Interfering; D051906:Smad7 Protein; C497807:Smad7 protein, mouse; D016212:Transforming Growth Factor beta; D008156:Luciferases