Abstract
Filamins are large dimeric F-actin cross-linking proteins, crucial for the mechanosensitive properties of a number of cell types. Due to their interaction with a variety of different proteins, they exert important regulatory functions. However, in the human testis the role of filamins has been insufficiently explored. Immunohistochemical staining of human testis samples identified filamin A (FLNA) in spermatogonia and peritubular myoid cells. Investigation of different testicular tumor samples indicated that seminoma also express FLNA. Moreover, mass spectrometric analyses identified FLNA as one of the most abundant proteins in human seminoma TCam-2 cells. We therefore focused on FLNA in TCam-2 cells, and identified by co-immunoprecipitation LAD1, RUVBL1 and DAZAP1, in addition to several cytoskeletal proteins, as interactors of FLNA. To study the role of FLNA in TCam-2 cells, we generated FLNA-deficient cells using the CRISPR/Cas9 system. Loss of FLNA causes an irregular arrangement of the actin cytoskeleton and mechanical instability, impaired adhesive properties and disturbed migratory behavior. Furthermore, transcriptional activity of typical stem cell factors is increased in the absence of FLNA. In summary, our data suggest that FLNA is crucially involved in balancing stem cell characteristics and invasive properties in human seminoma cells and possibly human testicular germ cells.
Highlights
Filamins (FLNs) are among the best investigated actin-binding proteins, and widely distributed in vertebrate and non-vertebrate cells [1,2,3]
We found that filamin A (FLNA) and FLNB are enriched in spermatogonial cells and in certain testicular germ cell tumors
Immunohistochemical analysis of human testicular biopsies using specific antibodies directed against FLNA or FLNB revealed that the two filamin isoforms have partly overlapping regions of localization
Summary
Filamins (FLNs) are among the best investigated actin-binding proteins, and widely distributed in vertebrate and non-vertebrate cells [1,2,3]. The human FLN family consists of three isoforms: FLNA, FLNB and FLNC. FLNA and FLNB are ubiquitously expressed, while expression of FLNC is more specific to skeletal and cardiac muscle [4]. Low expression of FLNC was reported for some non-muscle cells [5]. All FLNs display a similar structure, consisting of an N-terminal actin-binding domain, which contains two calponin homology domains, followed by a flexible rod segment composed of. All three FLN isoforms share a high sequence similarity and show, in some cell types, comparable expression patterns [8], implying that FLN isoforms share overlapping functions or can compensate for each other
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