Biology subjects

Khokha, M.

Publications and source records attributed to Khokha, M..

4 recordsLinked to original sources

bioRxiv2021

Kap-β2/Transportin mediates β-catenin nuclear transport in Wnt signaling

Wnt signaling is essential for many aspects of embryonic development including the formation of the primary embryonic axis. In addition, excessive Wnt signaling drives multiple diseases including cancer highlighting its importance for disease pathogenesis. {beta}-catenin is a key effector in this pathway that translocates into the nucleus and activates Wnt responsive genes. However, due to our lack of understanding of {beta}-catenin nuclear transport, therapeutic modulation of Wnt signaling has been challenging. Here, we took an unconventional approach to address this long-standing question by exploiting a heterologous model system, the budding yeast Saccharomyces cerevisiae, which contains a conserved nuclear transport machinery. In contrast to prior work, we demonstrate that {beta}-catenin accumulates in the nucleus in a Ran dependent manner, suggesting the use of a nuclear transport receptor (NTR). Indeed, a systematic and conditional inhibition of NTRs revealed that only Kap104, the orthologue of Kap-{beta}2/Transportin-1 (TNPO1), was required for {beta}-catenin nuclear import. We further demonstrate direct binding between TNPO1 and {beta}-catenin that is mediated by a conserved amino acid sequence that resembles a PY NLS. Finally, using Xenopus secondary axis and TCF/LEF reporter assays, we demonstrate that our results in yeast can be directly translated to vertebrates. By elucidating the NLS in {beta}-catenin and its cognate NTR, our study provides new therapeutic targets for a host of human diseases caused by excessive Wnt signaling. Indeed, we demonstrate that a small chimeric peptide designed to target TNPO1 can reduce Wnt signaling as a first step towards therapeutics.

developmental biology↗

bioRxiv2019

Novel truncating mutations in CTNND1 cause a dominant craniofacial and cardiac syndrome

AbstractCTNND1 encodes the p120-catenin (p120) protein, which has a wide range of functions, including the maintenance of cell-cell junctions, regulation of the epithelial-mesenchymal transition and transcriptional signaling. Due to advances in next generation sequencing, CTNND1 has been implicated in human diseases including cleft palate and blepharocheilodontic syndrome (BCD) albeit only recently. In this study, we identify eight novel protein-truncating variants, six de novo, in thirteen participants presenting with craniofacial dysmorphisms including cleft palate and hypodontia, as well as congenital cardiac anomalies, limb dysmorphologies and neurodevelopmental disorders. Using conditional deletions in mice as well as CRISPR/Cas9 approaches to target CTNND1 in Xenopus, we identified a subset of phenotypes that can be linked to p120-catenin in epithelial integrity and turnover, and additional phenotypes that suggest mesenchymal roles of CTNND1. We propose that CTNND1 variants have a wider developmental role than previously described, and that variations in this gene underlie not only cleft palate and BCD but may be expanded to a broader velocardiofacial-like syndrome.

genetics↗

bioRxiv2019

Differential turnover of Nup188 controls its levels at centrosomes and role in centriole duplication

NUP188 encodes a scaffold component of the nuclear pore complex (NPC) and has been implicated as a congenital heart disease gene through an ill-defined function at centrioles. Here, we explore the mechanisms that physically and functionally segregate Nup188 between the pericentriolar material (PCM) and NPCs throughout the cell cycle. Pulse-chase fluorescent labeling approaches indicate that Nup188 populates centrosomes with newly synthesized protein that does not exchange with NPCs even after mitotic NPC breakdown. In addition, the steady-state level of Nup188 at centrosomes is controlled by the sensitivity of the PCM pool, but not the NPC pool, to proteasomal degradation. Proximity-labeling and super-resolution microscopy supports that Nup188 interacts with components of PCM including Cep192 and the centriolar satellite component, PCM1. Consistent with this, Nup188 plays a role in centriole duplication at or upstream of Sas6 loading. Together, our data establish Nup188 as a functional component of PCM and potentially provides insight into the pathogenesis of congenital heart disease.

cell biology↗

bioRxiv2018

The Heterotaxy Gene, CCDC11, is Essential for Cytokinesis and Cell-Cell Adhesion Via RhoA Regulation

Mutations in CCDC11 have been identified in multiple patients with heterotaxy (Htx), a disorder of left-right (LR) patterning of the internal organs. In Xenopus, depletion of Ccdc11 causes defects in LR patterning, recapitulating the patient phenotype. Upon Ccdc11 depletion, normally monociliated cells of the Left-Right Organizer (LRO) exhibit multiple cilia per cell. Unexpectedly, we found that Ccdc11 is necessary for successful cytokinesis, and the multiciliation observed in Ccdc11-depleted cells was due to failed cytokinesis. Furthermore, CCDC11 depletion alters cell-cell adhesion with reduction in junctional localization of adhesion molecules. The small GTPase RhoA is critical for cytokinesis and cell-cell adhesion. Because the CCDC11 depletion phenotypes are reminiscent of RhoA loss of function, we investigated a possible connection to regulation of RhoA signaling. We demonstrate that CCDC11 is localized to the cytokinetic contractile ring overlapping with RhoA during cytokinesis and regulates total RhoA protein levels. Our results suggest that CCDC11 connects cytokinesis and LR patterning via RhoA regulation, providing a potential mechanism for heterotaxy disease pathogenesis.

cell biology↗