61 Oral - Cell Division and Cell Growth
Friday April 08, 8:45 AM - 9:00 AM

Authors:
Hala Zein-Sabatto ¹; Li Jin ²; Simon L. Bullock ²; Dorothy Lerit ¹

Affiliations:
1) Emory University Atlanta, GA, USA; 2) MRC Laboratory of Molecular Biology Cambridge, UK

Keywords:
d. centrosome; e. intracellular transport

The centrosome is a multi-functional organelle that plays a key role in nucleating and organizing microtubules, facilitating ciliogenesis, and organizing the bipolar mitotic spindle during cell division. A protein matrix known as the pericentriolar material (PCM) surrounds the centrosome and regulates centrosomal function. Various mRNAs also concentrate around the centrosome, but their functional significance is not yet fully understood. Our lab and others have identified centrocortin (cen) mRNA as forming micron-scale RNA granules near the centrosome in a cell cycle-dependent manner. We also showed localization of cen mRNA at the centrosomes is needed for error-free mitosis. However, how cen mRNA localizes to the centrosome is still unknown. Preliminary data from immunofluorescent imaging combined with smFISH reveals cen mRNA decorates astral microtubules. Further, biochemical studies show that Cen protein, a component of the cen mRNA granule, interacts with the dynein motor complex. Taken together, these data suggest that cen mRNA is transported to the centrosome via dynein-directed trafficking along microtubules. To test this hypothesis, cen mRNA localization was quantified in CRISPR-edited Drosophila embryos designed to disrupt a predicted dynein light intermediate chain binding site within the cen sequence. Similarly, cen mRNA localization was quantified in embryos of classical dynein heavy chain transheterozygous mutants. Moreover, the effect of destabilizing the microtubule network on cen granule localization to the centrosome was analyzed. Inhibiting the interaction of cen mRNA with dynein and microtubules resulted in a decrease in cen mRNA granules localized to the centrosome, consistent with a role for dynein in RNA trafficking to this site. Furthermore, fly lines harboring N- and C-terminal truncations of the cen transcript were made to identify the minimal sequence required for cen mRNA localization to the centrosome. This work provides insight into the dynamic localization of cen mRNA through active intracellular transport.