Biology subjects

Sagadiev, S.

Publications and source records attributed to Sagadiev, S..

4 recordsLinked to original sources

bioRxiv2025

The Lipid-binding PX Domain of RRC-1 (ARHGAP32/33) is Required for Optimal Assembly and Function of Integrin Adhesion Complexes at the Muscle Cell Boundary in C. elegans

Integrin adhesion complexes (IACs) are a network of many proteins that serve as anchors of the cell to the extracellular matrix (ECM). In muscle, IACs located at costameres, also serve to transmit the force of muscle contraction to the outside of the cell. We have reported that IACs, which are found at the bases of dense bodies and M-lines, and at muscle cell boundaries (MCB) in C. elegans muscle, require the RacGEF PIX-1 for their proper assembly or maintenance. We have reported that a RacGAP for the PIX pathway is RRC-1, is in a complex with PIX-1, and that RRC-1 is required for assembly or maintenance of IACs at MCBs. Our previous studies suggested that RRC-1 might be associated with the muscle cell membrane, and here we present evidence that this occurs via its PX domain, a domain that is known to bind to membrane phosphoinositides (PIPs). We predict the existence of a PX domain based on bioinformatic analysis and AlphaFold3, which includes conserved residues characteristic of most PX domains and a PIP binding site. This region of RRC-1 binds to phosphoinositides in vitro. Analysis of a nematode strain that has an in-frame deletion of the PX domain, indicates that normal localization of RRC-1 to the MCB requires both its PX domain and the PIX scaffold protein GIT-1. Lastly, we show that the overexpression of the full length RRC-1, but not RRC-1 with an in-frame deletion of its PX domain, results in reduced accumulation of IAC components and reduced whole animal movement. Our study highlights the importance of RRC-1s lipid interactions at the cell membrane for proper assembly and function of IACs in C. elegans muscle. Article SummaryIntegrin adhesion complexes (IACs) facilitate the transmission of force of muscle contraction to the outside of the cell. IACs, found at the bases of dense bodies and M-lines, and at muscle cell boundaries (MCB) in C. elegans muscle, require the PIX-1 (a RacGEF) signaling pathway for their assembly or maintenance. A RacGAP for the PIX pathway is RRC-1. Here, we show that RRC-1 has a PX domain that binds to membrane phosphoinositides. We also show that normal localization of RRC-1 to the MCB requires both its PX domain and the PIX scaffolding protein GIT-1.

cell biology↗

bioRxiv2025

IPMK-1 (inositol phosphate multikinase) is required for optimal integrin adhesion complex assembly in C. elegans muscle

Integrin adhesion complexes (IACs) are a network of many proteins including the transmembrane protein integrin that anchor cells to the extracellular matrix (ECM). In C. elegans muscle IACs exist at the bases of dense bodies and M-lines, and at the boundaries of adjacent muscle cells (MCBs). We have reported that the proper assembly of IACs requires the Rac GEF PIX-1 and members of the PIX-1 pathway. Here we report that further investigation into the original Million Mutation Project strain that led us to uncover the role of PIX-1 in muscle, revealed a genetic enhancer of the PIX-1 phenotype. Genetic mapping shows that the enhancer is a recessive mutation in a single gene, ipmk-1. ipmk-1, encodes inositol phosphate multikinase and converts PIP2 to PIP3, IP3 to IP4, and IP4 to IP5. Loss of function of pix-1 results in reduced accumulation of IAC proteins at the MCB, and ipmk-1; pix-1 double mutants additionally display a large gap between adjacent muscle cells. ipmk-1 similarly enhances other members of the PIX pathway including PAK-1, and the RacGAP RRC-1. Lack of ipmk-1 itself results in abnormal clumping of IAC components at the MCB, decreased whole animal locomotion, and mild sarcomere disorganization. Analysis of ipmk-1; ipp-5 double mutants suggest that the MCB defect of ipmk-1 is due to decreased PIP3 in the cell membrane rather than the decreased IP3 and Ca2+ signaling. Overall, these studies provide in vivo evidence, specifically in muscle, that the level of PIP3 and PIP2 influence the proper assembly of IACs. Article SummaryC. elegans muscle have integrin adhesion complexes (IACs) at dense bodies, M-lines, and boundaries between adjacent muscle cells (MCBs). The Benian lab reported that assembly of these IACs requires the PIX-1 pathway. Here they report ipmk-1 is a genetic enhancer of the PIX pathway. ipmk-1encodes inositol phosphate multikinase converting PIP2 to PIP3. pix-1 shows reduced accumulation of IAC components; ipmk-1; pix-1 double mutants additionally have large gaps between muscle cells. ipmk-1 mutants display abnormal clumping of IAC components. This is the first report of the importance of these lipids in assembly of IACs in vivo, and in particular, in muscle.

cell biology↗

bioRxiv2021

A screening pipeline identifies a broad-spectrum inhibitor of bacterial AB toxins with cross protection against influenza A virus H1N1 and SARS-CoV-2

A challenge for the development of host-targeted anti-infectives against a large spectrum of AB-like toxin-producing bacteria encompasses the identification of chemical compounds corrupting toxin transport through both endolysosomal and retrograde pathways. Here, we performed a high-throughput screening of small chemical compounds blocking active Rac1 proteasomal degradation triggered by the Cytotoxic Necrotizing Factor-1 (CNF1) toxin, followed by orthogonal screens against two AB toxins hijacking defined endolysosomal (Diphtheria toxin) or retrograde (Shiga-like toxin 1) pathways to intoxicate cells. This led to the identification of the molecule N-(3,3-diphenylpropyl)-1-propyl-4-piperidinamine, referred to as C910. This compound induces the swelling of EEA1-positive early endosomes, in absence of PIKfyve kinase inhibition, and disturbs the trafficking of CNF1 and the B-subunit of Shiga toxin along the endolysosomal or retrograde pathways, respectively. Together, we show that C910 protects cells against 8 bacterial AB toxins including large clostridial glucosylating toxins from Clostridium difficile. Of interest, C910 also reduced viral infection in vitro including influenza A virus subtype H1N1 and SARS-CoV-2. Moreover, parenteral administration of C910 to the mice resulted in its accumulation in lung tissues and reduced lethal influenza infection.

microbiology↗

bioRxiv2021

Transcriptomic analysis of pathways associated with alpha(v) integrin-related non-canonical autophagy in human B cells

Autophagy proteins have been linked with development of immune-mediated diseases including lupus, but the mechanisms for this are unclear. We have previously shown that non-canonical autophagy induced by v-integrins regulates B cell activation by viral and self-antigens in mice. Here we investigated the involvement of this pathway in B cells from human tissue. Our data revealed that autophagy is specifically induced in germinal-center and memory B cell sub-populations from human tonsil and spleen. Transcriptomic analysis showed that induction of autophagy is related to unique aspects of activated B cells such as mitochondrial metabolism. To understand the function of non-canonical autophagy in B cells, we used CRISPR-mediated knockdown of autophagy genes. Integrating data from primary B cells and knockout cells we found that v-integrin-related non-canonical autophagy limits activation of specific pathways while promoting others. These data provide new mechanistic links for autophagy and immune dysregulation in diseases such as lupus.

immunology↗