Dataset Information

Transforming growth factor-?1 (TGF-?1) regulates cell junction restructuring via Smad-mediated repression and clathrin-mediated endocytosis of nectin-like molecule 2 (Necl-2).

ABSTRACT: Nectin-like molecule-2 (Necl-2), a junction molecule, is exclusively expressed by spermatogenic cells. It mediates homophilic interaction between germ cells and heterophilic interaction between Sertoli and germ cells. Knockout studies have shown that loss of Necl-2 causes male infertility, suggesting Necl-2-based cell adhesion is crucial for spermatogenesis. Transforming growth factor-?s (TGF-?s) are crucial for regulating cell junction restructuring that are required for spermatogenesis. In the present study, we aim to investigate the mechanism on how TGF-?1 regulates Necl-2 expression to achieve timely junction restructuring in the seminiferous epithelium during spermatogenesis. We have demonstrated that TGF-?1 reduces Necl-2 mRNA and protein levels at both transcriptional and post-translational levels. Using inhibitor and clathrin shRNA, we have revealed that TGF-?1 induces Necl-2 protein degradation via clathrin-dependent endocytosis. Endocytosis assays further confirmed that TGF-?1 accelerates the internalization of Necl-2 protein to cytosol. Immunofluorescence staining also revealed that TGF-?1 effectively removes Necl-2 from cell-cell interface. In addition, TGF-?1 reduces Necl-2 mRNA via down-regulating Necl-2 promoter activity. Mutational studies coupled with knockdown experiments have shown that TGF-?1-induced Necl-2 repression requires activation of Smad proteins. EMSA and ChIP assays further confirmed that TGF-?1 promotes the binding of Smad proteins onto MyoD and CCAATa motifs in vitro and in vivo. Taken together, TGF-?1 is a potent cytokine that provides an effective mechanism in controlling Necl-2 expression in the testis via Smad-dependent gene repression and clathrin-mediated endocytosis.

SUBMITTER: Gao Y

PROVIDER: S-EPMC3669379 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Transforming growth factor-β1 (TGF-β1) regulates cell junction restructuring via Smad-mediated repression and clathrin-mediated endocytosis of nectin-like molecule 2 (Necl-2).

Gao Ying Y Lui Wing-Yee WY

PloS one 20130531 5

Nectin-like molecule-2 (Necl-2), a junction molecule, is exclusively expressed by spermatogenic cells. It mediates homophilic interaction between germ cells and heterophilic interaction between Sertoli and germ cells. Knockout studies have shown that loss of Necl-2 causes male infertility, suggesting Necl-2-based cell adhesion is crucial for spermatogenesis. Transforming growth factor-βs (TGF-βs) are crucial for regulating cell junction restructuring that are required for spermatogenesis. In the ...[more]

PMID: 23741316

Similar Datasets

Project description:BackgroundThe notion that AP-2 clathrin adaptor is an essential component of an endocytic clathrin coat appears to conflict with recent observations that substantial AP-2 depletion, using RNA interference with synthesis of AP-2 subunits, fails to block uptake of certain ligands known to internalize through a clathrin-based pathway.Methodology/principal findingsWe report here the use of in vivo imaging data obtained by spinning-disk confocal microscopy to study the formation of clathrin-coated structures at the plasma membranes of BSC1 and HeLa cells depleted by RNAi of the clathrin adaptor, AP-2. Very few clathrin coats continue to assemble after AP-2 knockdown. Moreover, there is a total absence of clathrin-containing structures completely lacking AP-2 while all the remaining coats still contain a small amount of AP-2. These observations suggest that AP-2 is essential for endocytic coated-pit and coated-vesicle formation. We also find that AP-2 knockdown strongly inhibits light-density lipoprotein (LDL) receptor-mediated endocytosis, as long as cells are maintained in complete serum and at 37 degrees C. If cells are first incubated with LDL at 4 degrees C, followed by warming, there is little or no decrease in LDL uptake with respect to control cells. LDL uptake at 37 degrees C is also not affected in AP-2 depleted cells first deprived of LDL by incubation with either serum-starved or LDL-starved cells for 24 hr. The LDL-deprived cells display a significant increase in endocytic structures enriched on deeply invaginated tubes that contain LDL and we suggest that under this condition of stress, LDL might enter through this alternative pathway.Conclusions/significanceThese results suggest that AP-2 is essential for endocytic clathrin coated-pit and coated-vesicle formation. They also indicate that under normal conditions, functional endocytic clathrin coated pits are required for LDL internalization. We also show that under certain conditions of stress, cells can upregulate alternative endocytic structures with the potential to provide compensatory trafficking pathways.

Dataset Information

Transforming growth factor-?1 (TGF-?1) regulates cell junction restructuring via Smad-mediated repression and clathrin-mediated endocytosis of nectin-like molecule 2 (Necl-2).

Publications

Transforming growth factor-β1 (TGF-β1) regulates cell junction restructuring via Smad-mediated repression and clathrin-mediated endocytosis of nectin-like molecule 2 (Necl-2).

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets