Molecular characterization and functional analysis of septin 8a in rohu (Labeo rohita): insights into immune regulation and developmental roles revealed by CRISPR/Cas9-mediated F0 mutagenesis.

Authors

Lopamudra Parida, Rajeeb K Swain, Pramoda Kumar Sahoo

Published in

Fish & shellfish immunology. Pages 111706. Sep 04, 2026. Epub Sep 04, 2026.

Abstract

Septins are conserved GTP-binding cytoskeletal proteins that regulate cytokinesis, membrane dynamics, and innate immunity. Here, we report the first molecular and functional characterization of septin 8a in rohu (Labeo rohita), an economically important Indian major carp. The full-length septin 8a cDNA encodes a 420-amino acid protein containing conserved GTP-binding motifs (G1-G5) belonging to the SEPT6 subgroup with a predicted molecular weight of ∼48.98 kDa, hydrophilic character and intracellular localization consistent with cytoskeletal scaffolding. Developmental expression analysis revealed peak transcript abundance at fertilization, suggesting maternal deposition and an early embryogenic role. Tissue profiling showed the highest constitutive expression in gills, kidney, and spleen. The challenge experiments with Aeromonas hydrophila, Argulus siamensis, and the viral mimic poly I:C demonstrated a broad pathogen-induced upregulation, particularly in immune tissues. Administration of generated recombinant septin 8a in vivo activated pro-inflammatory cytokines, antimicrobial enzymes, antioxidant genes, and the MX gene, and conferred 55% relative percent survival against A. hydrophila challenge. Indirect ELISA revealed progressive increase in protein levels within liver and kidney tissues during infection. CRISPR/Cas9-mediated F0 mutagenesis in zebrafish caused developmental delay, morphological abnormalities, and a survival rate of 26%, emphasizing its role during early development. Successful mutagenesis in rohu embryos validated cross-species sgRNA efficacy. Following bacterial challenge, exon 2-edited F0 crispants showed the highest mortality (80%) relative to wild-type controls (53.33%), whereas the mortality in exon 4-edited fish did not differ from controls. These findings establish septin 8a as a multifunctional immune regulator and developmental factor with potential applications in aquaculture disease management.

PMID:
42697518
Bibliographic data and abstract were imported from PubMed on 05 Sep 2026.

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