Abbi's first author publication accepted at Molecular Biology of the Cell!

by Heidi Hehnly in ,

Congrats to lead author and Hehnly Lab graduate student Abrar (Abbi) Aljiboury on the acceptance of her study titled “Pericentriolar matrix (PCM) integrity relies on cenexin and Polo-Like Kinase (PLK)1” at Molecular Biology of the Cell. This work was a collaborative study between Hehnly Lab post baccalaureates Amra Mujcic and Erin Curtis, undergraduates Denise Magny and Thomas Cammerino, graduate student Yiling Lan, the Blatt imaging center manager Mike Bates, Hehnly Lab manager Judy Freshour, and Biology faculty member Yasir Ahmed-Braimeh. This study examined PLK1 activity and its association with maintaining the functional and physical properties of the centrosome's pericentriolar matrix (PCM). Here, Abbi and colleagues use a multimodal approach of human cells (HeLa), zebrafish embryos, and phylogenic analysis to test the role of a PLK1 binding protein, cenexin, in regulating the PCM. Their studies identify that cenexin is required for tempering microtubule nucleation by maintaining PCM cohesion in a PLK1 dependent manner. PCM architecture in cenexin-depleted zebrafish embryos was rescued with wild-type human cenexin, but not with a C-terminal cenexin mutant (S796A) deficient in PLK1 binding. They propose a model where cenexin's C-terminus acts in a conserved manner in eukaryotes, excluding nematodes and arthropods, to sequester PLK1 that limits PCM substrate phosphorylation events required for PCM cohesion.

Cenexin is needed for PCM cohesion. (A) Metaphase HeLa cells mitotic centrosomes labeled for centrosome markers: centrin, cenexin, Cep192, Pericentrin, Cep215 and γ-tubulin (Fire LUT) and MT marker, α-tubulin (grey). Control shRNA (top) and cenexin shRNA (bottom) treated cells shown. Scale bar, 5 μm. (B-F) Representative scatter plots depicting two-dimensional areas (μm2) of centrin (B), Cep192 (C), Pericentrin (D), Cep215 (E) and γ-tubulin (F) in control and cenexin shRNA treated metaphase cells. Mean with 95% confidence intervals shown. Unpaired, two-tailed Student’s t-tests, n.s. not significant, ***p<0.001, ****p<0.0001. (G) Control shRNA (top) and cenexin shRNA (bottom) metaphase cell projection. Centrin (grey), Cep215 (magenta) and DNA (DAPI, cyan) shown. Insets magnified 3x from G’ and G”. Scale bar, 5 μm. (H) Model depicting representative centrosome protein outline from a single representative mitotic centrosome reflecting changes resulting from cenexin-loss. Mean 2-dimensional areas (μm2) ±SD are provided.

Check out new study from graduate student Nikhila Krishnan et al.

by Heidi Hehnly in

Title: Rab11 endosomes and Pericentrin coordinate centrosome movement during pre-abscission in vivo

The story can be found here.

We found that conserved between the zebrafish embryo and human cells, the oldest centrosome moves in a Rab11-dependent manner towards the cytokinetic bridge sometimes followed by the youngest. Rab11-endosomes are organized in a Rab11-GTP dependent manner at the mother centriole during pre-abscission, with Rab11 endosomes at the oldest centrosome being more mobile compared with the youngest. The GTPase activity of Rab11 is necessary for the centrosome protein, Pericentrin, to be enriched at the centrosome. Reduction in Pericentrin expression or optogenetic disruption of Rab11-endosome function inhibited both centrosome movement towards the cytokinetic bridge and abscission, resulting in daughter cells prone to being binucleated and/or having supernumerary centrosomes. These studies suggest that Rab11-endosomes contribute to centrosome function during pre-abscission by regulating Pericentrin organization resulting in appropriate centrosome movement towards the cytokinetic bridge and subsequent abscission.

Differences in mitotic centrosome movement towards the cytokinetic bridge during pre-abscission between zebrafish embryos and human cells.(A) Zebrafish embryo (5 h post fertilization) with centrin-GFP (gray) and PLK1-mCh (cyan). Scale bar, 50 μm. (a’) Inset of dividing cell in (A). Time-lapse of centrin-GFP (inverted grays, top panel; grays, bottom panel) and PLK1-mCh (cyan). Video 1 Pink arrow, centrosome. Orange arrow, midbody. Dashed lines, cell boundaries. Scale bar, 10 μm. (B) Model depicting centrosome (green) movement towards the cytokinetic bridge in dividing cells within the Kupffer’s vesicle (KV) during its development. Cyan, Nucleus. Purple, Midbody. Orange, Lumen. Dark lines, KV membranes. (C) Time-lapse of a dividing cell within the KV. KV cell membranes marked with Sox17:GFP-CAAX (gray). Cyan star, rosette center. Cyan arrow, centrosome. Dashed lines, cell boundaries. Scale Bar, 10 μm. (c’) Dividing cell depicted with PLK1-mCh (fire LUT). Cyan arrow, centrosome. (D) A KV pre-abscising cell fixed and immunostained for ZO-1 (gray), γ-tubulin (cyan) and DNA (DAPI, blue). Yellow arrow, centrosome. Dashed lines, cell boundaries. Scale bar, 10 μm. (E) Number of centrosomes per pre-abscising cell with bridge directed centrosome movement calculated as both centrosomes (2 centrosomes), only one centrosome (1 centrosome) and neither centrosome (0 centrosomes) moved shown as a violin plot with median (orange) and quartiles (dark dotted lines). Two-tailed t test between Centrin-GFP and DsRed-PACT in Human (HeLa) cells (pink background). n > 10 cells across n > 3 experiments n.s. not significant. One-way ANOVA across zebrafish epiboly cells and KV cells (green background), n.s. not significant. n > 10 cells across n > 2 embryos. One-way ANOVA, across all columns, **P < 0.01. Two-tailed t test between Human (HeLa) cells and zebrafish (Epiboly, KV) cells, **P < 0.01. n-values.