Figure 1.
Experimental design and integrative omics workflow.
A schematic diagram of the strategy used to access germ cell and Sertoli cell secretomes and to highlight potential protein-protein interactions. BTB: blood testis barrier; GC: germ cells; LC: Leydig cells; PC: peritubular cells; SC: Sertoli cells; Spg: spermatogonia; pSpc: pachytene spermatocytes; rSpt: round spermatids; eSPT: elongated spermatids.
Figure 2.
Details of datasets and the methods used to reconstruct dialog between Sertoli and germ cells.
(A) Conversion of the rat and ram UniProt identifiers (UPIDs) into mouse Entrez Gene identifiers (EGIDs). (B) Definition of the germ cell and Sertoli cell transcriptomes (PSIDs: Probeset identifiers) (C) Selection of loci (mouse Entrez gene IDs) encoding secreted or potentially secreted proteins (RSIDs: RefSeq identifiers UPIDs: UniProt identifiers; ENSIDs: Ensembl identifiers) (D) Selection of genes encoding proteins associated with a “plasma membrane” and/or “cell surface” location. (E) Selection of proteins secreted by one type of cell (germ or Sertoli cell) and interacting with membrane proteins of the other type of cell from BioGRID, HPRD, IntAct, MINT and NCBI databases.
Figure 3.
Integration of “omics” data to establish a network of molecular interactions between germ cells and Sertoli cells mediated by the TF.
This integrated network focuses on proteins produced and secreted by germ cells (GCs) or Sertoli cells (SCs) and interacting with membrane proteins of the other type of cell. Nodes symbolizing GC-secreted and Sertoli cell-secreted factors are indicated in light green and purple, respectively, whereas GC-membrane and Sertoli cell-membrane proteins are represented by red and light blue nodes, respectively.
Figure 4.
In situ detection of APOH/CDC42 and APP/NGFR interactions in the rat testis by Duolink PLA.
(A, E) Abundant PLA signals (red dots) were detected in the seminiferous tubules of rat testis when specific primary antibodies were used, reflecting the close proximity of APOH/CDC42 or APP/NGFR close proximity. (B, C, F, G) Negative controls, with only one primary antibody for the targeted protein-protein interaction. (D, H) Quantification of PLA signals in testis sections for APOH/CDC42 (** Student's t-test p = 0.0016) or for APP/NGFR (Student's t-test p = 0.16). Scale bars = 50 µm. A nonspecific background nuclear signal was observed for a few tubule sections (see B).