Small RNA and mRNA expression profiling during axolotl forelimb regeneration
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ABSTRACT: Previous studies of appendage regeneration in the axolotl have shown that multiple genetic programs are modulated through regulatory factors. MicroRNAs are short highly conserved non-coding genes that suppress expression of target genes and thereby control multiple genetic programs. Given their important regulatory roles and evolutionary conservation, we hypothesize that microRNAs define a conserved genetic regulatory circuit important for appendage regeneration. We characterized microRNA expression during Axolotl forelimb regeneration using small RNA sequencing. The same samples were assayed for mRNA expression using mRNA sequencing. Small RNA and mRNA gene expression profiling during 0, 3, 6 and 14 days post amputation.
Project description:Previous studies of vertebrate appendage regeneration have shown that multiple genetic programs are moduled through regulatory factors. MicroRNAs are short highly conserved non-coding genes that suppress expression of target genes and thereby control multiple genetic programs. Given their important regulatory roles and evolutionary conservation, we hypothesize that microRNAs define a conserved genetic regulatory circuit important for appendage regeneration. We characterized microRNA expression during Polypterus senegalus (bichir) pectoral fin regeneration using small RNA sequencing. The same samples were assayed for mRNA expression using mRNA sequencing. Small RNA and mRNA gene expression profiling during 0, 3, 7 and 14 days post amputation.
Project description:Previous studies of zebrafish caudal fin regeneration have shown that multiple genetic programs are moduled through regulatory factors. MicroRNAs are short highly conserved non-coding genes that suppress expression of target genes and thereby control multiple genetic programs. Given their important regulatory roles and evolutionary conservation, we hypothesize that microRNAs define a conserved genetic regulatory circuit important for appendage regeneration. We characterized microRNA expression during zebrafish caudal fin regeneration using small RNA sequencing. The stages of caudal fin regeneration were assayed for mRNA expression using mRNA sequencing. Small RNA and mRNA gene expression profiling during 0 and 4 days post amputation.
Project description:The goal of this study is to compare gene expression changes in retinal degenerate Royal College of Surgeons (RCS) rats following an injection of human neural progenitor cells (hNPCs) using RNA-seq, as compared to RCS rats receiving a sham surgery and wild-type Long Evans rats. These changes may lead to understanding of the therapeutic potential of hNPCs in inducing photoreceptor survival and visual function preservation. RNA-seq from wild-type Long Evans rats, retinal degenerate Royal College of Surgeons (RCS) rats, and RCS rats following transplantation of human neural progenitor cells (hNPCs)
Project description:The fate of doubled genes, from allopolyploid or autopolyploid origin, is controlled at multiple levels within the central dogma: gene loss or silencing, neo- and/or sub functionalization, inter genomic transfer, allele dominance/co-dominance, differences in transcription/translation efficiency, post translational modifications… These regulatory processes through evolution have caused a plethora of genotype x environment interactions displayed in the modern day phenotypes. The study of non-model crops is challenging but solutions are emerging. More and more, one gets insight into the tolerance mechanisms of a specific genotype. By integrating transcriptomics into our proteomic data, we studied the genetic diversity of an allopolyploid ABB banana, a tolerant genotype, and compared it to two different sensitive AAA genotypes. The root growth of the ABB cultivar was 60 % higher under mild osmotic stress. 234,000 spectra were aligned and quantified, resulting in 2,753 identified root proteins. 383 gene loci displayed genotype specific differential expression whereof 252 showed at least one Single Amino Acid Polymorphism (SAAP). The homeoallelic contribution was assessed using transcriptome read alignment, thus revealing each allele contribution at the RNA level. This provides insight in the structure and the organization of the triploid genome. In the ABB cultivar, allele expressions are supposed to follow a 1/3 and 2/3 pattern. We found that many genes deviated from this expectation and we show that 32 gene loci even displayed a 100% read preference for the allele that was unique for the ABB tolerant genotype , suggesting that the presence of unique alleles and homoelog expression bias is correlated to the observed phenotype.
Project description:Ribosome profiling data reports on the distribution of translating ribosomes, at steady-state, with codon-level resolution. We present a robust method to extract codon translation rates and protein synthesis rates from these data, and identify causal features associated with elongation and translation efficiency in physiological conditions in yeast. We show that neither elongation rate nor translational efficiency is improved by experimental manipulation of the abundance or body sequence of the rare AGG tRNA. Deletion of three of the four copies of the heavily used ACA tRNA shows a modest efficiency decrease that could be explained by other rate-reducing signals at gene start. This suggests that correlation between codon bias and efficiency arises as selection for codons to utilize translation machinery efficiently in highly translated genes. We also show a correlation between efficiency and RNA structure calculated both computationally and from recent structure probing data, as well as the Kozak initiation motif, which may comprise a mechanism to regulate initiation. We test whether tRNA abundance affects elongation or translation efficiency by changing the tRNA levels through deletion or over expression and measuring the ribosomal dwell time at each codon using a robust statistical method that accounts for flow conservation.
Project description:DEAD-box RNA helicases eIF4A and Ded1 are believed to promote translation initiation by resolving mRNA secondary structures that impede ribosome attachment at the mRNA 5’ end or subsequent scanning of the 5’UTR, but whether they perform distinct functions or act redundantly in vivo is poorly understood. We compared the effects of mutations in Ded1 or eIF4A on global translational efficiencies (TEs) in yeast by ribosome footprint profiling. Despite similar reductions in bulk translation, inactivation of a cold-sensitive Ded1 mutant substantially reduced the TEs of >600 mRNAs, whereas inactivation of a temperature-sensitive eIF4A mutant yielded <40 similarly impaired mRNAs. The broader requirement for Ded1 did not reflect more pervasive secondary structures at low temperature, as inactivation of temperature-sensitive and cold-sensitive ded1 mutants gave highly correlated results. Interestingly, Ded1-dependent mRNAs exhibit greater than average 5’UTR length and propensity for secondary structure, implicating Ded1 in scanning though structured 5' UTRs. Reporter assays confirmed that cap- distal stem-loop insertions increase dependence on Ded1 but not eIF4A for efficient translation. While only a small fraction of mRNAs is strongly dependent on eIF4A, this dependence is significantly correlated with requirements for Ded1 and 5’UTR features characteristic of Ded1- dependent mRNAs. Our findings suggest that Ded1 is critically required to promote scanning through secondary structures within 5’UTRs; and while eIF4A cooperates with Ded1 in this function, it also promotes a step of initiation common to virtually all yeast mRNAs. We compared the effects of mutations in Ded1 or eIF4A on global translational efficiencies (TEs) in yeast by ribosome footprint profiling.The study includes 32 samples, comprised of 16 mRNA-Seq samples and 16 ribosome footprint profiling samples, derived from biological replicates of 3 mutant strains, ded1-cs, ded1-ts and tif1-ts, and the corresponding wild-type strains. The tif1-ts mutant and its wild-type counterpart were analyzed at 30°C and 37°C.
Project description:Immunocytochemical studies revealed that dG9a moves into nucleus after cycle 8 and appears to regulate gene expression by di-methylating H3K9 from cycle 8 to cycle 11. To determine which genes are regulated by dG9a during cycles 8-11, we examined mRNA levels by performing RNA-sequence analysis using early embryos (0-2 h after egg laying) of dG9a null mutant and wild type as a control mRNA profiles of about 0-2h-old embryos of wild type (CantonS) and dG9a-depleted (dG9aRG5) strain
Project description:Brown adipose (BAT) development and function are unaffected by the absence of miR-193b/365-1 We refer to wild-type mice (+/+) as WT and to mice lacking the mir-193b (-/-) as Null mice.
Project description:The plasma cell transcription factor XBP1 is critical for terminal differentiation of B cells into plasma cells but has no known role at earlier stages of B-cell development. Here we show that XBP1 is not only important during early B-cell development and for survival of pre-B cells but also protects pre-B ALL cells. Among pre-B ALL subset, XBP1 was hypomethylated and highest expressed in the Ph+ ALL subset. Cre-mediated deletion of XBP1 in a mouse model of Ph+ ALL compromised proliferation and viability and prolonged survival of leukemia-bearing mice. Interestingly, XBP1 expression levels were positively transcriptionally regulated by STAT5 and negatively by BACH2 and BCL6. High XBP1 expression in high risk ALL patients at the time of diagnosis predicted poor outcome in two clinical trials. Clinically, small-molecule inhibition of IRE1-dependent XBP1-activation caused cell death of patient-derived pre-B ALL cells and affected leukemia-initiation in transplant recipient mice. Collectively, these studies identify XBP1 as an important survival factor and as a potential therapeutic target to overcome drug-resistance in pre-B ALL. Genome-wide profiling of mRNA levels in p210 transduced murine Xbp1 fl/+ pre-B cells with ERT2 (XE.1,2,3) and Cre- ERT2 M-BM- (XC.1,2,3).
Project description:SILAC labeled human kidney cells (293 cells) or bat kidney cells (PakiT03cells)were infected with Hendra virus for 8 or 24 hours and compared to uninfected control cells. Protein identification and quantitation relied on a combination of Uniprot lists of proteins and Proteomics Informed by Transcriptomics (PIT) analysis whereby RNA extracted from the same samples was deep sequenced and the sequencing data was used to construct mRNA from which possible ORFS were inferred and used as a search space by MaxQuant.