Biomedical subjects

Donglin Liu

Publications and source records attributed to Donglin Liu.

5 recordsLinked to original sources

PubMed2006

Systematic variation in mRNA 3'-processing signals during mouse spermatogenesis.

Gene expression and processing during mouse male germ cell maturation (spermatogenesis) is highly specialized. Previous reports have suggested that there is a high incidence of alternative 3'-processing in male germ cell mRNAs, including reduced usage of the canonical polyadenylation signal, AAUAAA. We used EST libraries generated from mouse testicular cells to identify 3'-processing sites used at various stages of spermatogenesis (spermatogonia, spermatocytes and round spermatids) and testicular somatic Sertoli cells. We assessed differences in 3'-processing characteristics in the testicular samples, compared to control sets of widely used 3'-processing sites. Using a new method for comparison of degenerate regulatory elements between sequence samples, we identified significant changes in the use of putative 3'-processing regulatory sequence elements in all spermatogenic cell types. In addition, we observed a trend towards truncated 3'-untranslated regions (3'-UTRs), with the most significant differences apparent in round spermatids. In contrast, Sertoli cells displayed a much smaller trend towards 3'-UTR truncation and no significant difference in 3'-processing regulatory sequences. Finally, we identified a number of genes encoding mRNAs that were specifically subject to alternative 3'-processing during meiosis and postmeiotic development. Our results highlight developmental differences in polyadenylation site choice and in the elements that likely control them during spermatogenesis.

3' Untranslated Regions↗

PubMed2006

Retinoic acid regulation of eye and testis-specific transcripts within a complex locus.

We previously used a yeast-based enhancer trap to identify a strong, retinoic acid response element (RARE). We have now characterized testis and eye transcripts that are adjacent to this regulatory element. Bioinformatics analysis of expressed sequence tag (EST) clones and RNase protection, reverse transcription-PCR, and Northern blot assays indicate that these two RNAs are transcribed from the same locus on opposite template strands. This positions the RARE upstream of the testis transcript and downstream of the eye transcript. Additionally, these two RNAs are embedded within the third intron of the 329kbp gene that encodes the Zinc Finger and BTB domain containing 7C protein (Zbtb7C). We present evidence indicating that the testis transcript is expressed primarily in spermatocytes and/or early round spermatids. Furthermore, our analyses of transcript levels in eyes and testes isolated from vitamin A deficient mice or from mice with defects in retinoid storage or signaling indicate that retinoids are required for expression in vivo.

Journal Article↗

PubMed2006

Quantitative comparison of EST libraries requires compensation for systematic biases in cDNA generation.

BACKGROUND: Publicly accessible EST libraries contain valuable information that can be utilized for studies of tissue-specific gene expression and processing of individual genes. This information is, however, confounded by multiple systematic effects arising from the procedures used to generate these libraries. RESULTS: We used alignment of ESTs against a reference set of transcripts to estimate the size distributions of the cDNA inserts and sampled mRNA transcripts in individual EST libraries and show how these measurements can be used to inform quantitative comparisons of libraries. While significant attention has been paid to the effects of normalization and substraction, we also find significant biases in transcript sampling introduced by the combined procedures of reverse transcription and selection of cDNA clones for sequencing. Using examples drawn from studies of mRNA 3'-processing (cleavage and polyadenylation), we demonstrate effects of the transcript sampling bias, and provide a method for identifying libraries that can be safely compared without bias. All data sets, supplemental data, and software are available at our supplemental web site. CONCLUSION: The biases we characterize in the transcript sampling of EST libraries represent a significant and heretofore under-appreciated source of false positive candidates for tissue-, cell type-, or developmental stage-specific activity or processing of genes. Uncorrected, quantitative comparison of dissimilar EST libraries will likely result in the identification of statistically significant, but biologically meaningless changes.

Algorithms↗

PubMed2005

PACdb: PolyA Cleavage Site and 3'-UTR Database.

UNLABELLED: The PolyA Cleavage Site and 3'-UTR Database (PACdb) is a web-accessible database that catalogs putative 3'-processing sites and 3'-UTR sequences for multiple organisms. Sites have been identified primarily via expressed sequence tag-genome alignments, enabling delineation of both the specificities and heterogeneity of 3'-processing events. AVAILABILITY: By web browser or CGI: PACdb: http://harlequin.jax.org/pacdb/; AtPACdb: http://harlequin.jax.org/atpacdb/. SUPPLEMENTARY INFORMATION: Available online at http://harlequin.jax.org/pacdb/supplemental.php.

3' Untranslated Regions↗

PubMed2004

Conservation of Bmp2 post-transcriptional regulatory mechanisms.

Bone morphogenetic protein (BMP) orthologs from diverse species like flies and humans are functionally interchangeable and play key roles in fundamental processes such as dorso-ventral axis formation in metazoans. Because both transcriptional and post-transcriptional mechanisms play central roles in modulating developmental protein levels, we have analyzed the 3'-untranslated region (3'UTR) of the Bmp 2 gene. This 3'UTR is unusually long and is alternatively polyadenylated. Mouse, human, and dog mRNAs are 83-87% identical within this region. A 265-nucleotide sequence, conserved between mammals, birds, frogs, and fish, is present in Bmp2 but not Bmp4. The ability of AmphiBMP2/4, a chordate ortholog to Bmp2 and Bmp4, to align with this sequence suggests that its function may have been lost in Bmp4. Activation of reporter genes by the conserved region acts by a post-transcriptional mechanism. Mouse, human, chick, and zebrafish Bmp2 synthetic RNAs decay rapidly in extracts from cells not expressing Bmp2. In contrast, these RNAs are relatively stable in extracts from Bmp2-expressing cells. Thus, Bmp2 RNA half-lives in vitro correlate with natural Bmp2 mRNA levels. The fact that non-murine RNAs interact appropriately with the mouse decay machinery suggests that the function of these cis-regulatory regions has been conserved for 450 million years since the fish and tetrapod lineages diverged. Overall, our results suggest that the Bmp2 3'UTR contains essential regulatory elements that act post-transcriptionally.

3' Untranslated Regions↗