Kindlin-3 splicing variants mark stress granulopoiesis 2261219

Introduction Mutations in the FERMT3 gene cause Leukocyte Adhesion Deficiency type III (LAD-III), a rare immunodeficiency characterized by recurrent infections. FERMT3 encodes kindlin-3, a protein essential for integrin activation, and a longer splice variant that differs by the insertion of four amino acids–Ile-Pro-Arg-Arg (IPRR)–within the pleckstrin homology (PH) domain (kindlin-3-IPRR). The expression pattern and function of the IPRR-containing isoform in neutrophil adhesion remain unclear. Methods Kindlin-3 knockout HL-60 cells and mouse hematopoietic stem cells (HSCs) were reconstituted with kindlin-3 short or kindlin-3-IPRR long isoforms by viral transduction. β2 integrin activation was assessed by an antibody reporter binding assay. Cell adhesion was measured in a flow chamber under flow or static conditions. FERMT3 splicing patterns in neutrophils from healthy donors and patients undergoing stress myelopoiesis induced by granulocyte colony-stimulating factor (G-CSF) or HSC transplantation were analyzed using RNA sequencing datasets. Results Kindlin-3-IPRR is functional in activating β2 integrins and supporting neutrophil adhesion. The IPRR insertion promotes kindlin-3 association with the plasma membrane. Analysis of RNA sequencing data reveals that the long kindlin-3-IPRR isoform is enriched in both hematopoietic stem cells and mature polymorphonuclear neutrophils (PMNs), suggesting a critical role in these populations. Kindlin-3-IPRR rescued the adhesion defect of kindlin-3 KO mouse HSCs. Notably, G-CSF treatment markedly increased expression of the long isoform in normal-density neutrophils (NDNs), whereas G-CSF—induced low-density neutrophils (LDNs) exhibited upregulated levels of the short isoform. Conclusion These findings suggest that stress conditions differentially regulate kindlin-3 isoform expression in human neutrophils. The kindlin-3-IPRR isoform is functional and may be necessary for NDN mobilization in response to G-CSF and could serve as a molecular signature of stress myelopoiesis.