The intestinal epithelium constitutes a crucial defense to the potentially life-threatening effects of gut microbiota. However, due to a complex underlying vasculature, hypoperfusion and resultant tissue ischemia pose a particular risk to function and integrity of the epithelium. The small ubiquitin-like modifier (SUMO) conjugation pathway critically regulates adaptive responses to metabolic stress and is of particular significance in the gut, as inducible knockout of the SUMO-conjugating enzyme Ubc9 results in rapid intestinal epithelial disintegration. Here we analyzed the pattern of individual SUMO isoforms in intestinal epithelium and investigated their roles in intestinal ischemia/reperfusion (I/R) damage. Immunostaining revealed that epithelial SUMO2/3 expression was almost exclusively limited to crypt epithelial nuclei in unchallenged mice. However, intestinal I/R or overexpression of Ubc9 caused a remarkable enhancement of epithelial SUMO2/3 staining along the crypt-villus axis. Unexpectedly, a similar pattern was found in SUMO1 knockout mice. Ubc9 transgenic mice, but also SUMO1 knockout mice were protected from I/R injury as evidenced by better preserved barrier function and blunted inflammatory responses. PCR array analysis of microdissected villus-tip epithelia revealed a specific epithelial contribution to reduced inflammatory responses in Ubc9 transgenic mice, as key chemotactic signaling molecules such as IL17A were significantly downregulated. Together, our data indicate a critical role particularly of the SUMO2/3 isoforms in modulating responses to I/R and provide the first evidence that SUMO1 deletion activates a compensatory process that protects from ischemic damage.
Ubc9 overexpression and SUMO1 deficiency blunt inflammation after intestinal ischemia/reperfusion.
Microarray analyses for the identification of differences in gene expression patterns have increased our understanding of the molecular genetic events in colorectal cancer.
A molecular signature for the prediction of recurrence in colorectal cancer.
Cellular differentiation requires both activation of target cell programs and repression of non-target cell programs. Transcriptional repressors such as RE1-silencing transcription factor (REST) and Hairy/Enhancer of Split (Hes) repress neuronal genes in non-neuronal cells. However, it is unknown whether transcriptional repressors of non-neuronal genes in neuronal precursors are required to specify neuronal fate during development. The Myt1 family of zinc finger transcription factors contributes to fibroblast to neuron reprogramming in vitro by repressing Notch signaling. Here, we show that ztf-11 (Zinc-finger Transcription Factor-11), the sole Caenorhabditis elegans Myt1 homolog, is required for neurogenesis in multiple neuronal lineages, including an in vivo developmental epithelial-to-neuronal transdifferentiation event. ztf-11 is exclusively expressed in all neuronal precursors with remarkable specificity at single cell resolution. Loss of ztf-11 leads to upregulation of non-neuronal genes and reduced neurogenesis. Ectopic expression of ztf-11 in epidermal lineages is sufficient to produce additional neurons. Our genetic and genomic experiments show that ZTF-11 indeed functions as a transcriptional repressor to suppress the activation of non-neuronal genes in neurons; however, it does not function via repression of Notch signaling. Instead, ZTF-11 binds to the MuvBco-repressor complex, which we show is also required for neurogenesis. These results dovetail with ability of Myt1l (Myt1-like) to drive neuronal transdifferentiation in vitro in vertebrate systems. Together, we identified an evolutionarily conserved mechanism to specify neuronal cell fate by repressing non-neuronal genes. Overall design: 4 biological replicates each under 2 experemental conditions (ztf-11 KD and negative control) were used for total of 8 samples
A Myt1 family transcription factor defines neuronal fate by repressing non-neuronal genes.
Specimen part, Cell line, Treatment, SubjectView Samples
Optically decodable beads link the identity of an analyte or sample to a measurement through an optical barcode, enabling libraries of biomolecules to be captured on beads in solution and decoded by fluorescence. This approach has been foundational to microarray, sequencing, and flow-based expression profiling technologies. We have combined microfluidics with optically decodable beads to link phenotypic analysis of living cells to sequencing. As a proof-of-concept, we applied this to demonstrate an accurate and scalable tool for connecting live cell imaging to single-cell RNA-Seq called Single Cell Optical Phenotyping and Expression (SCOPE-Seq). Overall design: Performed SCOPE-Seq on thousands of cells from two cell lines.
SCOPE-Seq: a scalable technology for linking live cell imaging and single-cell RNA sequencing.
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We sorted approx. 10000 neurons per sample from day one adult worms. We collected two wildtype samples and three thoc-5(wy822) mutant samples. Overall design: RNAseq of FACS-sorted C.elegans neurons from wildtype and thoc-5(wy822) mutant animals.
The THO Complex Coordinates Transcripts for Synapse Development and Dopamine Neuron Survival.
Specimen part, SubjectView Samples
Chemical induced pluripotent stem cells (CiPSCs) have been successfully achieved and may provide an alternative and attractive source for stem cell-based therapy. Sufficient telomere lengths are critical for unlimited self-renewal and genomic stability of pluripotent stem cells. Dynamics of telomere reprogramming and whether and how telomeres are sufficiently elongated in the CiPSCs have remained to be understood. We show that CiPSCs acquire telomere lengthening with increasing passages after clonal formation. Both telomerase activity and recombination-based mechanisms are involved in the telomere elongation. Telomere lengths strongly indicate the degree of reprogramming, pluripotency and differentiation capacity of CiPSCs. Nevertheless, telomere damage and shortening occur at late stage of lengthy induction, limiting CiPSC formation. Recombination mechanism is not activated during induction until CiPSC clonal formation and passages. We find that histone crotonylation induced by crotonic acid can activate two-cell genes including Zscan4, alleviate telomere damage and shortening during induction and promote CiPSC generation. Moreover, crotonylation decreases abundance of heterochromatic H3K9me3 and HP1a at subtelomeres and Zscan4 loci. Taken together, telomere rejuvenation links to reprogramming and pluripotency of CiPSCs. Crotonylation facilitates telomere maintenance and enhances chemical induced reprogramming to pluripotency. Overall design: We collected day 0, day 15, day 30, day 40 reprogramming cells and CiPSCs to detect telomeres dynamic. We collected day 20, day 24 and day 28 reprogramming cells with or without crotonic acid to do RNA-sequence experiment to analyze how CA can promote CiPSCs generation. We collected CiPS cell lines by three induction methods to analyze these cell lines transcriptome.
Dynamics of Telomere Rejuvenation during Chemical Induction to Pluripotent Stem Cells.
Specimen part, Cell line, SubjectView Samples
In birds and mammals, all mesoderm cells are generated from the primitive streak. Nascent mesoderm cells contain unique dorso-ventral (D/V) identities depending on their relative ingression position along the streak. Molecular mechanisms controlling this initial phase of mesoderm diversification are not well-understood. Using chick model, we generated high-quality transcriptomic datasets of different streak regions and analyzed their molecular heterogeneity.
Transcriptomic landscape of the primitive streak.
Specimen partView Samples
Blood was extracted from embryonic hearts at E4 and E6 and non-red blood was separated by density gradient centrifugation
Expression profiling of circulating non-red blood cells in embryonic blood.
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Various pluripotent stem (PS) cells can be isolated from early developing embryos in mouse. Among these, two kinds of PS cells were isolated from mouse blastocysts: conventional embryonic stem (ES) cells with domed morphology that are maintained with LIF and BMP for self-renewal, and FAB-ES cells with flat morphology that need bFGF, activinA and BIO for self-renewal. Here, we report a novel PS cell line from rat blastocysts, which is distinguishable from conventional ES cells but is morphologically similar to mouse epiblast stem cell (EpiSC) lines. We used microarrays to detail the global program of gene expression of rES and rPS.
The heterogeneity and dynamic equilibrium of rat embryonic stem cells.
Specimen partView Samples