To test the hypothesis that fast anterograde molecular motor proteins power the slow axonal transport of neurofilaments (NFs), we used homologous recombination to generate mice lacking the neuronal-specific conventional kinesin heavy chain, KIF5A. Because null KIF5A mutants die immediately after birth, a synapsin-promoted Cre-recombinase transgene was used to direct inactivation of KIF5A in neurons postnatally. Three fourths of such mutant mice exhibited seizures and death at around 3 wk of age; the remaining animals survived to 3 mo or longer. In young mutant animals, fast axonal transport appeared to be intact, but NF-H, as well as NF-M and NF-L, accumulated in the cell bodies of peripheral sensory neurons accompanied by a reduction in sensory axon caliber. Older animals also developed age-dependent sensory neuron degeneration, an accumulation of NF subunits in cell bodies and a reduction in axons, loss of large caliber axons, and hind limb paralysis. These data support the hypothesis that a conventional kinesin plays a role in the microtubule-dependent slow axonal transport of at least one cargo, the NF proteins.

Huntington's disease (HD) is an autosomal dominant neurodegenerative disease linked to a polyQ (polyglutamine) expansion in the huntingtin protein. Although general brain atrophy is found in HD patients, the striatum is the most severely affected region. Loss or mutant forms of huntingtin were reported to disrupt fast axonal transport in Drosophila, squid, and mice. However, previous work did not resolve whether mutant huntingtin affects global axonal transport or only a subset of cargoes, nor did it resolve whether striatal neurons are preferentially sensitive to huntingtin-mediated defects. We used amyloid precursor protein (APP)-yellow fluorescent protein and brain-derived neurotrophic factor (BDNF)-mCherry fusion proteins as markers for fast axonal transport when huntingtin is altered. We found that movement of APP and BDNF is impaired in striatal and hippocampal, but not cortical, neurons from presymptomatic homozygous mutant mice carrying 150Q huntingtin knock-in mutations. In addition, loss of huntingtin disrupts APP axonal transport, whereas overexpression of wild-type, but not mutant, huntingtin enhances APP transport in all three types of neurons tested. These data suggest that a loss of wild-type huntingtin function in fast axonal transport plays important roles in the development of cell-type-specific defects in HD.

The matrix (M) protein of vesicular stomatitis virus inhibits both nuclear import and export. Here, we demonstrate that this inhibitory property is conserved between the M proteins from two other vesiculoviruses, chandipura virus and spring viremia carp virus. All three M proteins completely block nuclear transport of spliced mRNA, small nuclear RNAs, and small nuclear ribonucleoproteins and slow the nuclear transport of many other cargoes. In all cases where transport was merely slowed by the M proteins, the chandipura virus M protein had the strongest inhibitory activity. When expressed in transfected HeLa cells, active M proteins displayed prominent association with the nuclear rim. Moreover, mutation of a conserved methionine abolished both the inhibitory activity and efficient targeting of the M proteins to the nuclear rim. We propose that all of the vesiculoviral M proteins associate with the same nuclear target, which is likely to be a component of the nuclear pore complex.nucleocytoplasmic transport ͉ vesiculoviruses ͉ RNA transport

MicroRNAs (miRNAs) play important roles in several neurobiological processes, including the development and progression of diseases. Previously, we identified that one specific miRNA, miR-196a, provides neuroprotective effects on Huntington's disease (HD), although the detailed mechanism is still unclear. Based on our bioinformatic analyses, we hypothesize miR-196a might offer neuroprotective functions through improving cytoskeletons of brain cells. Here, we show that miR-196a could enhance neuronal morphology, further ameliorating intracellular transport, synaptic plasticity, neuronal activity, and learning and memory abilities. Additionally, we found that miR-196a could suppress the expression of RAN binding protein 10 (RANBP10) through binding to its 3' untranslated region, and higher expression of RANBP10 exacerbates neuronal morphology and intracellular transport. Furthermore, miR-196a enhances neuronal morphology through suppressing RANBP10 and increasing the ability of β-tubulin polymerization. Most importantly, we observed higher expression of RANBP10 in the brains of HD transgenic mice, and higher expression of RANBP10 might exacerbate the pathological aggregates in HD. Taken together, we provide evidence that enhancement of neuronal morphology through RANBP10 is one of the neuroprotective mechanisms for miR-196a. Since miR-196a has also been reported in other neuronal diseases, this study might offer insights with regard to the therapeutic use of miR-196a in other neuronal diseases.

The SNT309 gene was identified via a mutation that causes lethality of cells in combination with a prp19 mutation. We showed previously that Snt309p is a component of the Prp19p-associated complex and that Snt309p, like Prp19p, is associated with the spliceosome immediately after or concomitantly with dissociation of U4 from the spliceosome. We show here that extracts prepared from the SNT309-deleted strain (⌬SNT309) were defective in splicing but could be complemented by addition of the purified Prp19p-associated complex. Isolation of the Prp19p-associated complex from ⌬SNT309 extracts indicated that the complex was destabilized in the absence of Snt309p and dissociated on affinity chromatography, suggesting a role of Snt309p in stabilization of the Prp19p-associated complex. Addition of the affinity-purified Prp19p-Snt309p binary complex to ⌬SNT309 extracts could reconstitute the Prp19p-associated complex. Genetic analysis further suggests that Snt309p plays a role in modulating interactions of Prp19p with other associated components to facilitate formation of the Prp19p-associated complex. A model for how Snt309p modulates such interactions is proposed.Splicing of pre-mRNA takes place on a multicomponent ribonucleoprotein particle called the spliceosome, which consists of five small nuclear RNAs (snRNAs) and a number of protein factors (see refs. 1-8 for reviews). Numerous protein factors involved in the splicing reaction have been identified in mammals and yeast. Some are intrinsic components of the spliceosome (9-11), whereas others associate with the spliceosome only transiently (12)(13)(14). Many of the protein factors are components of the sn ribonucleoprotein particles (see refs. 1 and 15-18 for reviews).Yeast genetics provides a powerful tool for identification of protein factors involved in the splicing reaction, as well as for studying interactions between these components. A large number of PRP (precursor RNA processing) genes that encode protein splicing factors have been identified by screening temperature-sensitive mutants defective in pre-mRNA splicing (19,20). Other genes were identified through genetic interactions with introns, PRP genes, or snRNA genes (21-27). Over 40 genes that encode protein factors of the splicing machinery have been identified, but the precise functions of these proteins are not well understood.The yeast PRP19 gene was among the genes identified in a screen for temperature-sensitive mutants defective in splicing (20). Biochemical characterizations indicated that the Prp19p protein is essential for the pre-mRNA splicing reaction in vitro.