Love, J. F. | bioRxiv Science

bioRxiv2024

Lysine Demethylase 4A is a Centrosome Associated Protein Required for Centrosome Integrity and Genomic Stability

Centrosomes play a fundamental role in nucleating and organizing microtubules in the cell and are vital for faithful chromosome segregation and maintenance of genomic stability. Loss of structural or functional integrity of centrosomes causes genomic instability and is a driver of oncogenesis. The lysine demethylase 4A (KDM4A) is an epigenetic eraser of chromatin methyl marks, which we show also localizes to the centrosome with single molecule resolution. We additionally discovered KDM4A demethylase enzymatic activity is required to maintain centrosome homeostasis, and is required for centrosome integrity, a new functionality unlinked to altered expression of genes regulating centrosome number. We find rather, that KDM4A interacts with both mother and daughter centriolar proteins to localize to the centrosome in all stages of mitosis. Loss of KDM4A results in supernumerary centrosomes and accrual of chromosome segregation errors including chromatin bridges and micronuclei, markers of genomic instability. In summary, these data highlight a novel role for an epigenetic eraser regulating centrosome integrity, mitotic fidelity, and genomic stability at the centrosome.

cell biology↗

bioRxiv2023

A hierarchical pathway for assembly of the distal appendages that organize primary cilia

Distal appendages are nine-fold symmetric blade-like structures attached to the distal end of the mother centriole. These structures are critical for formation of the primary cilium, by regulating at least four critical steps: ciliary vesicle recruitment, recruitment and initiation of intraflagellar transport (IFT), and removal of CP110. While specific proteins that localize to the distal appendages have been identified, how exactly each protein functions to achieve the multiple roles of the distal appendages is poorly understood. Here we comprehensively analyze known and newly discovered distal appendage proteins (CEP83, SCLT1, CEP164, TTBK2, FBF1, CEP89, KIZ, ANKRD26, PIDD1, LRRC45, NCS1, C3ORF14) for their precise localization, order of recruitment, and their roles in each step of cilia formation. Using CRISPR-Cas9 knockouts, we show that the order of the recruitment of the distal appendage proteins is highly interconnected and a more complex hierarchy. Our analysis highlights two protein modules, CEP83-SCLT1 and CEP164-TTBK2, as critical for structural assembly of distal appendages. Functional assay revealed that CEP89 selectively functions in RAB34+ ciliary vesicle recruitment, while deletion of the integral components, CEP83-SCLT1-CEP164-TTBK2, severely compromised all four steps of cilium formation. Collectively, our analyses provide a more comprehensive view of the organization and the function of the distal appendage, paving the way for molecular understanding of ciliary assembly.

cell biology↗