We describe Trans-ABySS, a de novo short-read transcriptome assembly and analysis pipeline that addresses variation in local read densities by assembling read substrings with varying stringencies and then merging the resulting contigs before analysis. Analyzing 7.4 gigabases of 50-base-pair paired-end Illumina reads from an adult mouse liver poly(A) RNA library, we identified known, new and alternative structures in expressed transcripts, and achieved high sensitivity and specificity relative to reference-based assembly methods.

The MAD-related (MADR) family of proteins are essential components in the signaling pathways of serine/threonine kinase receptors for the transforming growth factor beta (TGFbeta) superfamily. We demonstrate that MADR2 is specifically regulated by TGFbeta and not bone morphogenetic proteins. The gene for MADR2 was found to reside on chromosome 18q21, near DPC4, another MADR protein implicated in pancreatic cancer. Mutational analysis of MADR2 in sporadic tumors identified four missense mutations in colorectal carcinomas, two of which display a loss of heterozygosity. Biochemical and functional analysis of three of these demonstrates that the mutations are inactivating. These findings suggest that MADR2 is a tumor suppressor and that mutations acquired in colorectal carcinomas may function to disrupt TGFbeta signaling.

MAD-related (MADR) proteins are essential intracellular components of TGFbeta signaling pathways and are regulated by phosphorylation. Here, we demonstrate that MADR2 and not the related protein DPC4 transiently interacts with the TGFbeta receptor and is directly phosphorylated by the complex on C-terminal serines. Interaction of MADR2 with receptors and phosphorylation requires activation of receptor I by receptor II and is mediated by the receptor I kinase. Mutation of the phosphorylation sites generates a dominant negative MADR2 that blocks TGFbeta-dependent transcriptional responses, stably associates with receptors, and fails to accumulate in the nucleus in response to TGFbeta signaling. Thus, transient association and phosphorylation of MADR2 by the TGFbeta receptor is necessary for nuclear accumulation and initiation of signaling.

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