Biology subjects

Buster, D. W.

Publications and source records attributed to Buster, D. W..

4 recordsLinked to original sources

bioRxiv2025

Transient centrosome loss in cultured prostate epithelial cells induces chromosomal instability to produce an oncogenic genotype that correlates with poor clinical outcomes

Chromosomal instability (CIN) is a hallmark of prostate cancer that strongly correlates with metastatic burden and appears prominently in both primary cancer and metastatic disease. Low Gleason score primary prostate tumors display pervasive centrosome loss, a known mechanistic driver of CIN, that disrupts normal spindle assembly and increases mitotic errors. Previously, we found that transient depletion of centrosomes in immortalized, non-tumorigenic prostate epithelial cells (PrEC) induced a burst of CIN, generating cell lines capable of forming xenograft tumors. Here, we use a multi-omics approach to identify the oncogenic alterations caused by transient centrosome loss. By integrating genomic and transcriptomic data of the prostate lines, we identified a consensus set of focal copy-number variations (CNVs) induced by centrosome loss in cultured cells that are also detectable within a subset of samples from a prostate cancer patient cohort. Using this CNV signature, we were able to derive a unique transcriptomic signature from prostate cancer patient samples that showed strong predictive value for adverse clinical outcomes. In summary, our experimental system uses centrosome loss to promote a punctuated burst of genomic crisis that is characteristic of genome evolution during prostate cancer progression. Consequently, this prostate cancer model produced recurrent structural variations that are detectable in patient samples and associate with worse outcomes. Graphic Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=140 SRC="FIGDIR/small/653266v1_ufig1.gif" ALT="Figure 1"> View larger version (55K): org.highwire.dtl.DTLVardef@ac4c9org.highwire.dtl.DTLVardef@fd9961org.highwire.dtl.DTLVardef@160e20corg.highwire.dtl.DTLVardef@198bf0e_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2021

Polo-like kinase 4 homodimerization is not required for catalytic activation, autodestruction, or centriole assembly

Polo-like kinase 4 (Plk4) is the master-regulator of centriole assembly and cell cycle-dependent regulation of its activity maintains proper centrosome number. During most of the cell cycle, Plk4 levels are nearly undetectable due to its ability to autophosphorylate and trigger its own ubiquitin-mediated degradation. However, during mitotic exit, Plk4 forms a single aggregate on the centriole surface to stimulate centriole duplication. Whereas most Polo-like kinase family members are monomeric, Plk4 is unique because it forms homodimers. Notably, Plk4 trans-autophosphorylates a degron near its kinase domain, a critical step in autodestruction. While it is thought that the purpose of homodimerization is to promote trans-autophosphorylation, this has not been tested. Here, we generate separation-of-function Plk4 mutants that fail to dimerize, and we show that homodimerization is required to create a binding site for the Plk4 activator, Asterless. Surprisingly, Plk4 dimer mutants are catalytically active in cells, promote centriole assembly, and can trans-autophosphorylate by a process based its on concentration-dependent aggregation. Our findings implicate a concentration-dependent pathway of Plk4 activation that does not require Asterless binding or homodimerization. Lastly, we propose a model of Plk4 cell cycle regulation that utilizes both activation pathways - Asterless-dependent and aggregation-driven - to restrict centriole assembly to mother centrioles.

cell biology↗

bioRxiv2019

A molecular mechanism for the procentriole recruitment of Ana2

Centriole duplication begins with the assembly of a pre-procentriole at a single site on a mother centriole and proceeds with the hierarchical recruitment of a conserved set of proteins, including Polo-like kinase 4 (Plk4)/ZYG-1, Ana2/SAS-5/STIL, and the cartwheel protein Sas6. During assembly, Ana2/STIL stimulates Plk4 kinase activity, and in turn, Ana2/STILs C-terminus is phosphorylated, allowing it to bind and recruit Sas6. The assembly steps immediately preceding Sas6-loading appear clear, but the mechanism underlying the upstream pre-procentriole recruitment of Ana2/STIL is not. In contrast to proposed models of Ana2/STIL recruitment, we recently showed that Drosophila Ana2 targets procentrioles independent of Plk4-binding. Instead, Ana2 recruitment requires Plk4 phosphorylation of Ana2s N-terminus, but the mechanism explaining this process is unknown. Here, we show that the amyloid-like domain of Sas4, a centriole surface protein, binds Plk4 and Ana2, and facilitates phosphorylation of Ana2s N-terminus which increases Ana2s affinity for Sas4. Consequently, Ana2 accumulates at the procentriole to induce daughter centriole assembly.

cell biology↗

bioRxiv2017

An ordered pattern of Ana2 phosphorylation by Plk4 is required for centriole assembly

Polo-like kinase 4 (Plk4) initiates an early step in centriole assembly by phosphorylating Ana2/STIL, a structural component of the procentriole. Here, we show that Plk4 binding to the central coiled-coil (CC) of Ana2 is a conserved event, involving Polo-box 3 and a previously unidentified putative CC located adjacent to the kinase domain. Ana2 binding stimulates Plk4 kinase activity in vitro, and, in turn, is phosphorylated along its length. Previous studies showed that Plk4 phosphorylates the C-terminal STAN domain of Ana2/STIL, triggering binding and recruitment of the cartwheel protein Sas6 to the procentriole assembly site. However, the physiological relevance of N-terminal phosphorylation was unknown. We found that Plk4 first phosphorylates the extreme N-terminus of Ana2 which is critical for subsequent STAN domain modification. Phosphorylation of the central region then breaks the Plk4-Ana2 interaction. This phosphorylation pattern is important for centriole assembly and integrity because replacement of endogenous Ana2 with phospho-Ana2 mutants disrupts distinct steps in Ana2 function and inhibits centriole duplication.

cell biology↗