Project description:The human skeletal muscle transcriptome – sex differences, alternative splicing and tissue homogeneity assessed with RNA sequencing
Project description:Exon level expression analysis for the physiological aging study data set to analyze the effect of age on alternative splicing in different tissues and age groups of wild-type mice Analysis of the effect of age on alternative splicing (AS) using exon microarrays to interrogate the differential exon usage of the entire genome of aging wild-type male C57BL/6J mice (4- and 18-month-old) in five tissues (skin, skeletal muscle, bone, thymus, and white adipose tissue) and in an additional 28-month-old age group, which allowed for age-related AS analysis of the skin, skeletal muscle and bone tissues. We found AS genes with age in all tissue, we show that the number of AS genes increased with age and that AS genes across all tissues are involved in RNA processing. Note: This dataset is one of the 2 datasets in the overall study. An additional data set series is available with exon expression analysis of HGPS mice to analyze the effect of progerin expression on alternative splicing. The two datasets are linked together in the SuperSeries GSE67289. A link to the SuperSeries is available at the bottom of this page. 65 tissue samples from wild-type male C57BL/6J mice; from 5 different tissues (ventral skin, skeletal muscle, bone, muscle, and white adipose tissue) and from 3 different age groups: 4, 18 and 28 months (for skin skeletal muscle and bone ) and from 2 different age groups: 4 and 18 months (for ventral skin, skeletal muscle, bone, thymus and white adipose tissue)
Project description:The molecular mechanisms underlying the sex differences in human muscle morphology and function remain to be elucidated. The purpose of the study was to detect the sex differences in the skeletal muscle transcriptome in both the resting state and following anabolic stimuli, resistance exericse.
Project description:Hnrnpu is the largest member of the heterogenous nuclear ribonucleoprotein family of RNA binding proteins. Hnrnpu is involved in pre-mRNA alternative splicing regulation. We used high throughput sequencing to determine how Hnrnpu regulates skeletal muscle physiology.
Project description:We carried out the first analysis of alternative splicing complexity in human tissues using mRNA-Seq data. New splice junctions were detected in 20% of multiexon genes, many of which are tissue specific. By combining mRNA-Seq and EST-cDNA sequence data, we estimate that transcripts from 95% of multiexon genes undergo alternative splicing and that there are 100,000 intermediate- to high-abundance alternative splicing events in major human tissues. From a comparison with quantitative alternative splicing microarray profiling data, we also show that mRNA-Seq data provide reliable measurements for exon inclusion levels. Keywords: mRNA expression 32-nucleotide sequence reads from six human tissues including brain, cerebral cortex, heart, liver, lung and skeletal muscle.
Project description:Smooth muscle cells (SMCs) are important in a number of physiological systems and organs, including the cardiovascular system. The hallmark property of differentiated SMCs is the ability to contract, but contractile SMCs themselves show a range of phenotypes allowing prolonged tonic contraction in vascular smooth muscle or rapid phasic contraction in tissues such as bladder. Another distinctive characteristic, in contrast with terminally differentiated striated muscle cells, is that SMCs exhibit phenotypic plasticity. Vascular SMCs are able to modulate their phenotype along a continuum between a contractile phenotype, characteristic of healthy blood vessels, and a more proliferative âsyntheticâ phenotype, so-named for the enhanced synthesis and secretion of extracellular matrix components. Synthetic phenotype cells are found in a number of pathological situations such as atherosclerosis and arterial injury. We used mouse exon-junction (MJAY) arrays to gain insights into both the global contribution of alternative splicing events in re-shaping the transcriptome of dedifferentiating mouse aorta and bladder SMCs, and into the underlying regulatory mechanisms of the alternative splicing program. Affymetrix splice junction arrays (MJAY) were used to profile changes in both alternative splicing and transcript levels during the phenotypic modulation of smooth muscle cells when placed in culture. RNA extracted from intact aorta and bladder smooth muscle tissue was used for differentiated samples. For dedifferentiated, proliferative samples smooth muscle cells were enzymatically dispersed and grown in tissue culture for a week. Triplicate RNA samples were prepared from smooth muscle tissue of mouse aorta and bladder (differentiated) and from smooth muscle cells from each tissue cultured for 7 days (proliferative). The samples allowed comparison of alternative splicing (and other transcriptome) changes between differentiated and proliferative smooth muscle cell samples from two distinct types of smooth muscle cell, as well as allowing direct comparison of aorta (tonic smooth muscle) and bladder (phasic smooth muscle).
Project description:Myotonic dystrophes (DM), the most common adult muscular dystrophy, are the first recognized examples of RNA-mediated diseases in which expression of mutant RNAs containing expanded CUG or CCUG repeats interfere with the splicing of other mRNAs. Using whole-genome microarrays, we found that alternative splicing of the BIN1 mRNA is altered in DM skeletal muscle tissues, resulting in the expression of an inactive form of BIN1 deprived of phosphoinositide-binding and membrane-tubulating activities. BIN1 is involved in tubular invaginations of the plasma membrane and is essential for biogenesis of the muscle T-tubules, which are specialized skeletal muscle membrane structures essential to correct excitation-contraction (E-C) coupling. Mutations in the BIN1 gene cause centronuclear myopathy (CNM) that shares some histopathological features with DM, and both diseases are characterized by muscle weakness. Consistent with a loss-of-function of BIN1, muscle T-tubules were altered in DM patients, and membrane tubulation was restored upon expression of the correct splicing form of BIN1 in DM muscle cells. By deciphering the mechanism of BIN1 splicing mis-regulation we demonstrate that the splicing regulator, MBNL1, which is sequestered by expanded CUG and CCUG in DM, binds the BIN1 pre-mRNA and regulates directly its alternative splicing. Finally, reproducing BIN1 splicing alteration in mice is sufficient to reproduce the DM features of T-tubule alterations and muscle weakness. We propose that alteration of BIN1 alternative splicing regulation leads to muscle weakness, a predominant pathological feature of DM. Exon-Array analysis of control and CDM1 muscle primary cultures 10 days of differentiation
Project description:How species with similar repertoires of protein coding genes differ so dramatically at the phenotypic level is poorly understood. From comparing the transcriptomes of multiple organs from vertebrate species spanning ~350 million years of evolution, we observe significant differences in alternative splicing complexity between the main vertebrate lineages, with the highest complexity in the primate lineage. Moreover, within as little as six million years, the splicing profiles of physiologically-equivalent organs have diverged to the extent that they are more strongly related to the identity of a species than they are to organ type. Most vertebrate species-specific splicing patterns are governed by the highly variable use of a largely conserved cis-regulatory code. However, a smaller number of pronounced species-dependent splicing changes are predicted to remodel interactions involving factors acting at multiple steps in gene regulation. These events are expected to further contribute to the dramatic diversification of alternative splicing as well as to other gene regulatory changes that contribute to phenotypic differences among vertebrate species. mRNA profiles of several organs (brain, liver, kidney, heart, skeletal muscle) in multiple vertebrate species (mouse, chicken, lizard, frog, pufferfish) generated by deep sequencing using Illumina HiSeq
Project description:A number of microRNAs have been shown to regulate skeletal muscle development and differentiation. MicroRNA-222 is downregulated during myogenic differentiation and its overexpression leads to alteration of muscle differentiation process and specialized structures. By using RNA induced silencing complex (RISC) pulldown followed by RNA sequencing, combined with in silico microRNA target prediction, we have identified two new targets of microRNA-222 involved in the regulation of myogenic differentiation, Ahnak and Rbm24. Specifically, the RNA binding protein Rbm24 is a major regulator of muscle specific alternative splicing and its downregulation by microRNA-222 results in defective exon inclusion impairing the production of muscle-specific isoforms of Coro6, Fxr1 and NACA transcripts. Reconstitution of normal levels of Rbm24 in cells overexpressing microRNA-222 rescues muscle-specific splicing. In conclusion, we have identified a new function of microRNA-222 leading to alteration of myogenic differentiation at the level of alternative splicing, and we provide evidence that this effect is mediated by Rbm24 protein. We built linear models using 2 different experiments and two conditions (miR222 over expression (n=1) and control siRNA(n=2)) with the linear formula (~condition + experiment).