Figure 3.

Unmasking the functional role of Linc00513 in spermatogenesis. A) Luciferase reporter assay analysis of Linc00513’s effect on TGFBR1 expression. Co-transfection of Linc00513 with the TGFBR1 3’ UTR resulted in a significant increase in luciferase activity compared to TGFBR1 3’ UTR alone, suggesting that Linc00513 suppresses miR-7-mediated repression of TGFBR1; B) Western blot analysis of TGFBR1 protein levels in azoospermic testes. TGFBR1 protein expression was significantly down regulated in TCam-2 cell line compared to HS1 cell line, suggesting that miR-7 may be overactive in azoospermia; C) Western blot analysis of Linc00513 overexpression in TCam-2 cell line. Overexpression of Linc00513 in TCam-2 cells resulted in a significant increase in TGFBR1 protein expression, similar to the effect observed in HS1 cells; D) Western blot analysis of miR-7 overexpression in HS1 cells. Overexpression of miR-7 in HS1 cells resulted in a significant decrease in TGFBR1 protein expression.