Interactions and Phosphorylation-Dependent Roles of ARHGAP25 in Neutrophils
Sasvári Péter
Molecular Medicine
Dr. Várnai Péter
NET Zöld előadóterem
2025-12-10 12:15:00
Cellular and Molecular Physiology
Dr. Hunyady László
Dr. Csépányi-Kömi Roland
Dr. Bőgel Gábor
Dr. Pap Ádám
Dr. Sebestyén Anna
Dr. Ruisanchez Éva
Dr. Sveiczer Ákos
ARHGAP25 is a GTPase-activating protein playing a regulatory role in neutrophils and potentially other leukocytes. Nonetheless, many details remain unresolved, particularly regarding its intracellular behavior. This thesis aims to expand our current knowledge and uncover unexplored areas related to this protein.
First, we examined its phosphorylation-dependent functions in vivo by generating and transfecting overexpressed Ser-to-Ala mutants in PLB-985 cells and measuring their RAC levels, along with their F-actin content. The overexpression of the mutation-bearing ARHGAP25 proteins did not have a clear effect on RAC-activation, but the S379+380 phosphosites were deemed relevant for the regulation of filamentous actin levels.
To pinpoint protein interactions and pathways involved with ARHGAP25, we compiled a list of protein partners using human neutrophilic granulocytes as the starting material, yielding 76 proteins. Subsequent functional enrichment analysis identified several expected and novel cellular functions based on the list, proposing new perspectives to ARHGAP25-related research.
The results identified 14‑3‑3 members as candidates. In silico analysis identified eight potential interaction sites on ARHGAP25, and we demonstrated that prior phosphorylation of GST-ARHGAP25 facilitates its association with 14‑3‑3 proteins. This interaction may represent a novel mechanism for the regulation of ARHGAP25, though additional research is required to support this hypothesis.
The identification of four small G proteins (RAC2, RHOG, ARF4, and RAB27A) produced novel results. The following in silico studies corroborated the viability of the interaction between RHOG and ARHGAP25, associated via ARHGAP25's GAP domain, and the interaction was influenced by the activation cycle of the small GTPase.
