Identification of novel TDRD7 isoforms

Abstract

The aim of our study was to investigate the tudor domain-containing protein 7 (TDRD7) subcellular localization, which could be linked to diverse functions of this protein within the cell. Methods. In this study we employed cell imaging technique for detecting TDRD7 subcellular localization, Western blot analysis of HEK293 cell fractions with anti-TDRD7 monoclonal antibodies and bioinformatical search of possible TDRD7 isoforms in Uniprot, Ensemble, UCSC databases. Results. We have observed specific TDRD7-containing structures in cytoplasm as well as in the nucleus in HEK293 cells. The Western blot analysis of subcellular fractions (cytoplasm, mitochondria, nucleus) allowed us to detect three lower immunoreactive bands, with the aproximate molecular weight of 130, 110 and 60 kDa (we termed them as TDRD7b, TDRD7g and TDRD7d) and specific subcellular localization. The bioinformatical analysis of TDRD7 primary structure allowed us to determine two alternative transcripts from TDRD7 gene coding for proteins with calculated molecular weight of 130 and 60 kDa. Conclusion. The presented data demonstrate the existence at protein level of potential TDRD7 isoforms: TDRD7b, TDRD7g and TDRD7d. The expression profile of these splice variants and their role in cells remains to be elucidated.

Adaptor protein TDRD7 is co-localized with ribosomal protein S6 kinases S6K1 and S6K2 in cell lines of different tissue origin

“…Our recent studies with employment of immunofluorescentconfocal microscopy and anti-TDRD7 (E6) monoclonal antibody indicated the presence of TDRD7 not only in the cytoplasm, but in the nucleus as well. Especially strong TDRD7 positive signal was observed in the perinuclear zone and in the region of mitotic spindle formation [19].…”

“…Our previous studies allowed us to determine TDRD7 distribution in HEK293 cells [19]. Using anti-TDRD7 monoclonal antibodies (E6) we had demonstrated that a large fraction of this protein was localized in the nucleus and perinuclear region of HEK293 cells.…”

Early germline differentiation in bivalves: TDRD7 as a candidate investigational unit for Ruditapes philippinarum germ granule assembly

“…Indeed, the Tudor protein family is one of the most conserved groups of molecular determinants in germline specification (Fierro-Constaín et al 2017 ). The TUDOR domain is commonly found in a wide range of proteins that are involved in RNA metabolism and splicing, histone modification, DNA damage response, cell division, differentiation, genome stability and gametogenesis (Hosokawa et al 2007 ; Skorokhod et al 2011 ; Pek et al 2012 ). These determinants are usually organized in ribonucleoprotein complexes in the germ plasm of PGCs and/or nuage localised near nuclear pores (Fierro-Constaín et al 2017 ).…”

“…In other species, TDRD7 orthologues have been observed in cytoplasmic, mostly perinuclear positions. However, western blots of cellular fractions revealed the presence of a short isoform of human TDRD7 (60 kDa) in the nucleus, while longer isoforms were located in cytoplasmic and mitochondrial fractions (Skorokhod et al 2011 ). Moreover, Tanaka and colleagues ( 2011 ) observed the presence of TDRD7 in nuclei of mammal intermediate differentiating germ cells, i.e.…”

“…Ambiguous observations resulted from the western blot analysis. In previous works, western blots of subcellular fractions (cytoplasm, mitochondria, nucleus; Skorokhod et al 2011 ) and immunofluorescence (Hosokawa et al 2007 ; Patil et al 2014 ) allowed to investigate the localisation of TDRD7 in the mouse and the fruit fly. It was mostly found into specific structures in the cytoplasm: on the mitochondria outer membrane and in the perinuclear zone (Hosokawa et al 2007 ; Skorokhod et al 2011 ) and localised to the perinuclear nuage with Vasa (Patil et al 2014 ).…”