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Gary J. Bassell, Email: gary.bassell@emory.edu
R.P.H. designed plasmids and performed transfections, stable cell line creation, microscopy, and data analysis related to the centrosome recruitment assay, neurite fractionation, MEF fractionation, and MCP-Halo experiments. K.R.M. performed all experiments related to the split kinesin assay, immunofluorescence in primary neurons, and live neuronal imaging of MBNL granules. A.J.K. and L.K. contributed equally to this work. A.J.K. cloned full-length mScarlet-I kinesin plasmids and performed live imaging and data analysis of MBNL-kinesin co-transport in live neurons. L.K. performed co-immunoprecipitation experiments of MBNL1 with full-length kinesins and colocalization experiments with endogenous kinesin and MBNL1 in fixed cells. L.T.D. and T.S participated in plasmid construction and experimental design. D.P.B. and Z.L. performed experiments involving transfection and data analysis of centrosome recruitment with single MBNL1 ZnF mutants. K.L. assisted with study design. G.J.B and E.T.W. conceived of the project, supervised, and assisted in the design of all study components, and provided resources and funding. R.P.H., G.J.B., and E.T.W. wrote the paper.
We dedicate this paper posthumously to K.L., who was a graduate student at Emory University. We thank Gary Banker (Oregon Health and Science University) for providing kinesin tail and cargo-expressing plasmids used in the split kinesin assay and Andy Berglund (University at Albany) for providing MBNL1 plasmids with RIM and CM mutations. We thank Edouard Bertrand (Institut de Génétique Moléculaire de Montpellier) for the donation of a plasmid expressing a 45xMS2 array that was used to design the inducible RNA reporter. We also thank Maury Swanson (University of Florida) for donating MBNL1/2 double knockout fibroblasts, Chris Burge (Massachusetts Institute of Technology) for donating N2A and CAD cells, and Anita Corbitt (Emory University) for donating a plasmid containing the sequence for ZC3H14. An additional thanks to Arielle Valdez-Sinon for technical assistance with co-immunoprecipitation experiments, Kendra McKee for technical assistance to prepare RNAseq libraries, Hailey Olafson for RNA-seq data processing, and Chase Kelley for assistance with particle tracking analysis. We thank Belinda Pinto, Chase Kelley, and other members of the Wang and Bassell labs for insightful comments and suggestions. Finally, we thank the Emory University Microscopy Core and University of Florida Center for Neurogenetics for use of microscopy equipment in fixed and live cell imaging experiments. L.K. was supported by NIH NRSA training grant F31 NS117086. This work was supported by an NIH R01 NS114253 awarded to G.J.B. and E.T.W., and a Ben Barres Early Career Acceleration Grant from the Chan Zuckerberg Initiative awarded to E.T.W.
The authors declare no competing interests.
© The Author(s) 2023