SUN-134 CYTOSKELETON-ASSOCIATED PROTEIN 4 (CKAP4), A NEW PLAYER IN THE REGULATION OF MESANGIAL CELL PROLIFERATION

Abstract

Cytoskeleton-associated protein 4 (CKAP4) has recently attracted attention within cancer research, where its overexpression has been associated with increased lung, pancreatic and kidney cancer cell proliferation and a worsened prognosis. However, the potential role of CKAP4 in mesangioproliferative diseases such as IgA nephropathy (IgAN) and diabetic nephropathy (DN) has not previously been studied. The main growth factor known to induce proliferation in mesangial cells is PDGF, and the mesangial cells are the only glomerular cells known to express PDGF receptor beta (PDGFRβ). PDGF is normally secreted by the glomerular endothelial cells as well as the mesangial cells themselves resulting in a paracrine and autocrine regulation of mesangial cell proliferation. Increased PDGF levels in the glomeruli leads to mesangial proliferation as seen in DN and IgAN. Our hypothesis is that CKAP4 is involved in the regulation of mesangial proliferation. Expression of CKAP4 in glomeruli was investigated using immunofluorescence and immunoEM on human renal sections. CKAP4 was silenced using shRNA in primary human mesangial cells. Westernblot and masspectrometry was used to study protein expression and BrdU proliferation assay to determine proliferation. In order to understand protein interaction co-immunoprecipitation was used. Finally, two different mouse models of DN was used to investigate CKAP4 expression in disease. Immunohistochemistry as well as immunoEM showed expression of CKAP4 in glomerular mesangial cells and in the cell bodies of podocytes. Silencing CKAP4 resulted in an abated mitogenic response to the growth factor PDGF-BB, and further experiments revealed a significantly decreased expression of the PDGF receptor, both on RNA and protein level. Preliminary results from co-immunoprecipitation experiments indicate that PDGFRβ and CKAP4 interact. Analysis of the total protein content in the silenced cells using mass spectrometry showed only small or no changes in expression of other growth factor receptors such as EGFR and TGFbRs. In two different mouse models of DN CKAP4 was found to be significantly upregulated in later stages of disease with established mesangial expansion and other key features of the disease. We here report for the first time that CKAP4 is expressed in mesangial cells and that the expression of CKAP4 influences the cellular proliferative response to PDGF-BB and the expression of PDGFRβ. CKAP4 has previously been reported to interact with the EGF receptor on the cell surface of hepatocellular carcinomas and thereby influencing the growth of these cells. The EGF receptor and the PDGF receptor has several similarities and are tyrosine kinase receptors. We suggest that the interaction between the PDGF receptor and CKAP4 resembles the one of CKAP4 and EGFR. CKAP4 could be a potential mediator and target of mesangial proliferation in glomerular kidney disease.