Involvement of the fungal nuclear migration gene nudC human homolog in cell proliferation and mitotic spindle formation. Journal   Experimental cell research. Citation   Exp Cell Res. 273(1):73-84 Publication date   2002 Feb 1 Authors   Zhang MY Huang NN Clawson GA Osmani SA Pan W Xin P Razzaque MS Miller BA Investigators   Gary A. Clawson Weihua Pan Min-Ying Zhang Grant agencies   National Institute of Diabetes and Digestive and Kidney Diseases National Institute of General Medical Sciences National Center for Research Resources Grants   NIDDK DK 46778 NIDDK F32 DK09790-03 NIGMS GM-42564 NCRR M01 RR10732 MeSH headings   Cell Division Mitotic Spindle Apparatus Proteins MeSH qualifiers   physiology Abstract   Essential genes which are required for normal nuclear migration and play a role in developmental processes have been isolated from model genetic organisms. One such gene is nudC (nuclear distribution C), which is required for positioning nuclei in the cytoplasm of the filamentous fungus Aspergillus nidulans and for normal colony growth. This gene is highly conserved, structurally and functionally, throughout evolution and the human homolog, HnudC, has been cloned. To study the function of nudC in higher eukaryotic cells, HnudC was downregulated by developing triple ribozyme constructs, consisting of two cis-acting ribozymes which liberate an internal trans-acting ribozyme targeted to HnudC. Efficient cleavage sites in HnudC mRNA were identified using a library selection technique and HnudC-targeted internal ribozymes were cloned into a triple ribozyme cassette. Triple ribozyme constructs were subcloned into an ecdysone-inducible expression vector and stably transfected into human embryonic 293 cells. Muristerone A induced expression of the HnudC ribozyme and produced specific reduction of HnudC mRNA. Downregulation of HnudC mRNA resulted in significant inhibition of cell proliferation in clones expressing the HnudC-targeted triple ribozyme, which was not observed in uninduced cells or cells transfected with vector alone. In induced cultures, many mitotic cells demonstrated defects in spindle architecture during mitosis. The most common defect observed was multiple mitotic spindle poles rather than the expected bipolar structure. These data demonstrate the fundamental importance of HnudC in eukaryotic cell proliferation and a functional role for HnudC in spindle formation at mitosis. Medline ID   21655055