Characterization of FAM114A1: a novel podocyte cytoskeleton-associated protein upregulated in glomerular injury

Transcriptomic analysis of microdissected human glomeruli has suggested novel molecular signatures associated with membranous nephropathy (MN) by revealing several genes differentially upregulated in MN compared with other glomerular diseases. We focused on a novel protein, family with sequence similarity 114 member A1 (FAM114A1), that was identified as the top classifier gene in the dataset. To determine the localization of FAM114A1 within glomeruli, we performed immunofluorescence (IF) staining on normal human kidney specimens. The staining area was quantitated in human MN and rat passive Heymann nephritis (PHN). In addition, we analyzed the expression of FAM114A1 in cultured podocytes and C57BL/6N mice following lipopolysaccharide (LPS)-induced injury. In silico investigations were conducted to model the protein structure of FAM114A1. We knocked down FAM114A1 in cultured podocytes by siRNA transfection and conducted functional assays. To detect interacting proteins, an affinity pulldown assay was performed using FAM114A1-3XFLAG protein and human glomerular extract. IF studies demonstrated the majority of FAM114A1 staining localized to the primary and foot processes of podocytes. The expression of FAM114A1 was increased in human MN and rat PHN and with LPS-induced injury. In silico modeling revealed that FAM114A1 is an all-alpha protein with several conserved regions. In cultured podocytes, FAM114A1 colocalized with F-actin and focal adhesion molecules. Silencing FAM114A1 affected podocyte cytoskeletal development, podocyte cell migration, and cell attachment. Affinity pulldown screening revealed that FAM114A1 interacts with several cytoskeleton-associated proteins. These findings suggest that FAM114A1 is a novel podocyte cytoskeleton-associated protein whose expression is upregulated by glomerular injury.NEW & NOTEWORTHY Podocyte cytoskeletal proteins are crucial for podocyte integrity and maintenance of slit diaphragms as urinary filtration barriers. In this study, we focused on a novel protein, FAM114A1, that was the top classifier gene in MN in the gene expression study. We show that FAM114A1 is a podocyte-specific protein in the kidney and is upregulated in glomerular injury. FAM114A1 is associated with the podocyte cytoskeleton and silencing FAM114A1 affected podocyte cell morphology and functions.