Khan, R. B. | bioRxiv Science

bioRxiv2026

The KN domain of KANK proteins contains separable talin-binding and intramolecular interaction modules

KANK proteins link integrin adhesions to the cortical microtubule stabilising complex (CMSC) through interactions with the adhesion adaptor talin. However, how KANK proteins are regulated remains unclear. Here we show that the KN domain of KANK proteins contains separable regions that mediate talin binding and a conserved intramolecular interaction. Using fluorescence polarisation, NMR spectroscopy and structural analysis, we map an interaction between the N-terminal KN domain and the C-terminal ankyrin repeat domain and identify residues 60-68 of the KN domain as required for this intramolecular interaction. In contrast, the canonical LD motif within residues 30-60 mediates binding to talin. Deletion of residues 60-68 disrupts the intramolecular interaction while preserving talin binding, demonstrating that the KN domain contains distinct modules for talin engagement and intramolecular regulation. This regulatory architecture is conserved across the KANK family, although sequence variation modulates the strength of the intramolecular interaction. Together, these findings identify a modular organisation within the KANK KN domain that separates talin recognition from intramolecular regulation and is consistent with an autoinhibitory mechanism. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=122 SRC="FIGDIR/small/731086v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@e2dd22org.highwire.dtl.DTLVardef@240996org.highwire.dtl.DTLVardef@1f1926borg.highwire.dtl.DTLVardef@c3e523_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical Abstract.C_FLOATNO The KN domain of KANK proteins contains separable talin-binding and intramolecular interaction modules.The LD motif (blue) mediates binding to the talin R7 domain, whereas residues 60-68 (yellow) are required for interaction with the C-terminal ankyrin repeat domain. An AlphaFold model is shown as a structural interpretation of the intramolecular KN-ankyrin repeat interaction identified in this study. C_FIG

biochemistry↗

bioRxiv2025

A single factor for safer cellular rejuvenation

Ageing is a key driver of the major diseases afflicting the modern world. Slowing or reversing the ageing process would therefore drive significant and broad benefits to human health. Previously, the Yamanaka factors (OCT4, SOX2, KLF4, with or without c-MYC: OSK(M)) have been shown to rejuvenate cells based on accurate predictors of age known as epigenetic clocks. Unfortunately, OSK(M) induces dangerous pluripotency pathways, making it unsuitable for therapeutic use. To overcome this therapeutic barrier, we screened for novel factors by optimising directly for age reversal rather than for pluripotency. We trained a transcriptomic ageing clock, unhindered by the low throughput of bulk DNA methylation assays, to enable a screen of unprecedented scale and granularity. Our platform identified SB000, the first single gene intervention to rejuvenate cells from multiple germ layers with efficacy rivalling the Yamanaka factors. Cells rejuvenated by SB000 retain their somatic identity, without evidence of pluripotency or loss of function. These results reveal that decoupling pluripotency from cell rejuvenation does not remove the ability to rejuvenate multiple cell types. This discovery paves the way for cell rejuvenation therapeutics that can be broadly applied across age-driven diseases. HighlightsO_LISB000 drives multi-omic rejuvenation in human fibroblasts, as evidenced by substantial reversal of numerous epigenetic clocks, lowered single-cell transcriptomic age, and decreased senescence-associated gene expression. C_LIO_LIIn contrast to OSK(M), SB000 treatment maintains transcriptomic and functional measures of fibroblast identity without the activation of pluripotency. C_LIO_LISB000 rejuvenation generalises to keratinocytes, cells from another germ layer, with potency matching or surpassing OSK(M). C_LI

cell biology↗