Characterization of SKAP/kinastrin isoforms: the N-terminus defines tissue specificity and Pontin binding

Abstract

Small Kinetochore-Associated Protein (SKAP)/Kinastrin is a multifunctional protein with proposed roles in mitosis, apoptosis and cell migration. Exact mechanisms underlying its activities in these cellular processes are not completely understood. SKAP is predicted to have different isoforms, however, previous studies did not differentiate between them. Since distinct molecular architectures of protein isoforms often influence their localization and functions, this study aimed to examine the expression profile and functional differences between SKAP isoforms in human and mouse. Analyses of various human tissues and cells of different origin by RT-PCR, and by Western blotting and immunocytochemistry applying newly generated anti-SKAP monoclonal antibodies revealed that human SKAP exists in two protein isoforms: ubiquitously expressed SKAP16 and testis/sperm-specific SKAP1. In mouse, SKAP1 expression is detectable in testis at 4 weeks postnatally, when the first wave of spermatogenesis in mice is complete and the elongated spermatids are present in the testes. Furthermore, we identified Pontin as a new SKAP1 interaction partner. SKAP1 and Pontin co-localized in the flagellar region of human sperm suggesting a functional relevance for SKAP1-Pontin interaction in sperm motility. Since most previous studies on SKAP were performed with the testis-specific isoform SKAP1, our findings provide a new basis for future studies on the role of SKAP in both human somatic cells and male germ cells, including studies on male fertility.

Proteomic Landscape of Human Sperm in Patients with Different Spermatogenic Impairments

“…RUVBL1 functions as a DNA-dependent ATPase with ATP-dependent DNA helicase activity and is involved in several transcription complexes and histone modifications [ 61 , 62 , 63 ]. RUVBL1 interacts with the testis/sperm-specific Small Kinetochore-Associated Protein (SKAP1) and is located in the sperm flagellum [ 64 ]. In addition, RUVBL1 is a part of the R2TP complex, which acts as a co-chaperon together with the heat shock protein 90 (HSP90) [ 65 ].…”

“…HSP90 is also located in the sperm flagellum and is crucial to maintain male fertility. It was hypothesized that a co-chaperon complex is formed in sperm thereby contributing to the maturation of sperm [ 64 , 65 ]. The RUVBL1 protein was positively correlated with sperm count.…”