The cerebellum consists of a highly organized set of folia that are largely generated postnatally during expansion of the granule cell precursor (GCP)pool. Since the secreted factor sonic hedgehog (Shh) is expressed in Purkinje cells and functions as a GCP mitogen in vitro, it is possible that Shh influences foliation during cerebellum development by regulating the position and/or size of lobes. We studied how Shh and its transcriptional mediators,the Gli proteins, regulate GCP proliferation in vivo, and tested whether they influence foliation. We demonstrate that Shh expression correlates spatially and temporally with foliation. Expression of the Shh target gene Gli1 is also highest in the anterior medial cerebellum, but is restricted to proliferating GCPs and Bergmann glia. By contrast, Gli2is expressed uniformly in all cells in the developing cerebellum except Purkinje cells and Gli3 is broadly expressed along the anteroposterior axis. Whereas Gli mutants have a normal cerebellum, Gli2 mutants have greatly reduced foliation at birth and a decrease in GCPs. In a complementary study using transgenic mice, we show that overexpressing Shh in the normal domain does not grossly alter the basic foliation pattern, but does lead to prolonged proliferation of GCPs and an increase in the overall size of the cerebellum. Taken together, these studies demonstrate that positive Shh signaling through Gli2 is required to generate a sufficient number of GCPs for proper lobe growth.

SUMMARY Alternative intronic polyadenylation (IPA) can generate truncated protein isoforms with significantly altered functions. Here, we describe 31 dominant-negative, secreted variant isoforms of receptor tyrosine kinases (RTKs) that are produced by activation of intronic poly(A) sites. We show that blocking U1-snRNP can activate IPA, indicating a larger role for U1-snRNP in RNA surveillance. Moreover, we report the development of an antisense-based method to effectively and specifically activate expression of individual soluble decoy RTKs (sdRTKs) to alter signaling, with potential therapeutic implications. In particular, a quantitative switch from signal transducing full-length vascular endothelial growth factor receptor-2 (VEGFR2/KDR) to a dominant-negative sKDR results in a strong anti-angiogenic effect both on directly targeted cells and on naïve cells exposed to conditioned media, suggesting a role for this approach in interfering with angiogenic paracrine and autocrine loops.

Splicing factor hnRNPH drives an oncogenic splicing switch in gliomasThis study reveals two alternative splicing events that contribute to the development of glioma. HnRNPH is shown to control production of a pro-survival splice variant of the death-domain adaptor protein IG20-MADD and the motility-enhancing isoform of the RON receptor tyrosine kinase.

Public reporting burden for this collection of information is estimated to average 1 hour per response, including the time for reviewing instructions, searching existing data sources, gathering and maintaining the data needed, and completing and reviewing this collection of information. Send comments regarding this burden estimate or any other aspect of this collection of information, including suggestions for reducing this burden to Department of Defense, Washington Headquarters Services, Directorate for Information ERG, a member of the ETS transcription factor family, is frequently overexpressed in prostate cancer as a result of its fusion to the androgenresponsive Tmprss2 gene. Different genomic rearrangements and alternative splicing events around the junction region lead to multiple combination of Tmprss2:ERG fusion transcripts that correlate with different tumor aggressiveness, but their specific functions and biological activities are still unclear. The complexity of ERG expression pattern is compounded by the use of alternative promoters, splice sites, polyadenylation sites and translation initiation sites in both the native and fusion contexts. Our systematic characterization of native ERG and Tmprss2:ERG variants reveals that their different oncogenic potential is impacted by the status of the Ets domain and the configuration of the 5' UTR region. In particular, expression and activity of functional ERG and Tmprss2:ERG variants are influenced both by translation initiation signals within the different isoforms and by inhibitory upstream Open Reading Frames (uORF) in their 5' UTRs. Stable expression of ERG and Tmprss2:ERG variants promoted cell migration/invasion, induced a block of proliferation and induced a senescence-like state, suggesting a role for these variants in the prostate tumorigenesis process. In addition to Tmprss2:ERG fusion products, a group of related native ERG isoforms is also highly over-expressed in fusioncarrying prostate cancers, and share the same translation initiation site (in ERG exon 4) with the commonly observed Tmprss2 exon1 joined to ERG exon 4 (T1:E4) fusion-derived variant. Usage of this ATG can be preferentially down-regulated by directed antisense-based compounds, possibly representing the basis of a targeted approach that distinguishes between tumor-associated and normal ERG.