Cloning and embryonic expression patterns of the chicken CELF family (original) (raw)
Related papers
Journal of Molecular and Cellular Cardiology, 2009
During the transition from juvenile to adult life, the heart undergoes programmed remodeling at the levels of transcription and alternative splicing. Members of the CUG-BP and ETR-3-like factor (CELF) family have been implicated in driving developmental transitions in alternative splicing of cardiac transcripts during maturation of the heart. Here, we investigated the timing of the requirement for CELF activity in the postnatal heart using a previously described transgenic mouse model (MHC-CELFΔ). In MHC-CELFΔ mice, nuclear CELF activity has been disrupted specifically in the heart by cardiac-specific expression of a dominant negative CELF protein. Longitudinal analyses of two lines of MHC-CELFΔ mice with differing levels of dominant negative protein expression demonstrate that CELF splicing activity is required for healthy cardiac function during juvenile, but not adult, life. Cardiac function, chamber dilation, and heart size all recover with age in the mild line of MHC-CELFΔ mice without a loss of dominant negative protein expression or change in expression of endogenous CELF proteins or known CELF antagonists. This is the first example of a mouse model with genetically induced cardiomyopathy that spontaneously recovers without intervention. Our results suggest that CELF proteins are key players in the integrated gene expression program involved in postnatal cardiac remodeling and maturation.
Expression of cVg1 mRNA during chicken embryonic development
The Anatomical Record, 2003
Using degenerated PCR-primers to identify known and novel BMPs that are expressed in the developing chicken heart, we identified not only BMP2, -4, and -7 mRNA, but also the TGF superfamily member cVg1. The expression pattern of cVg1 mRNA was determined during chicken development from HH4 to HH44. In early developmental stages, cVg1 mRNA is expressed in the primitive streak, paraxial mesoderm, developing somites, and developing neural tube. Subsequently, cVg1 mRNA is expressed in the developing central and peripheral nervous system, retina, auditory vesicle, notochord, lung alveoli, and olfactory mucosa. In the heart, cVg1 is initially expressed through the linear heart tube, but becomes restricted to the forming chamber myocardium, in an expression domain similar to that of atrial natriuretic factor (ANF) mRNA. Anat Rec Part A 273A:603-608, 2003.
Molecular cloning of chicken Cecr2 and its expression during chicken embryo development
The International Journal of Developmental Biology, 2010
Cecr2 is a transcription factor involved in neurulation and chromatin remodeling. In the present study, the full length of the coding sequence of the chicken orthologue Cecr2 was obtained by RT-PCR. Sequence analysis and alignment showed that it contained an AT hook, as well as a bromodomain which was highly conserved among different species, consistent with its role in chromatin remodeling. The expression pattern of chicken Cecr2 was subsequently investigated during the development of the chicken embryo by in situ hybridization. In addition to its predominant expression in neural tissues during neurulation, Cecr2 was also found to be expressed in the developing somites and in the intermediate zone of the spinal cord, suggesting that it may play a role in somite and neuronal development.
CELF proteins regulate CFTR pre-mRNA splicing: essential role of the divergent domain of ETR-3
Nucleic Acids Research, 2010
Cystic fibrosis is a prominent genetic disease caused by mutations of the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Among the many disease-causing alterations are pre-mRNA splicing defects that can hamper mandatory exon inclusion. CFTR exon 9 splicing depends in part on a polymorphic UG(m)U(n) sequence at the end of intron 8, which can be bound by TDP-43, leading to partial exon 9 skipping. CELF proteins, like CUG-BP1 and ETR-3, can also bind UG repeats and regulate splicing. We show here that ETR-3, but not CUG-BP1, strongly stimulates exon 9 skipping, although both proteins bind efficiently to the same RNA motif as TDP-43 and with higher affinity. We further show that the skipping of this exon may be due to the functional antagonism between U2AF 65 and ETR-3 binding onto the polymorphic U or UG stretch, respectively. Importantly, we demonstrate that the divergent domain of ETR-3 is critical for CFTR exon 9 skipping, as shown by deletion and domain-swapping experiments. We propose a model whereby several RNA-binding events account for the complex regulation of CFTR exon 9 inclusion, with strikingly distinct activities of ETR-3 and CUG-BP1, related to the structure of their divergent domain.
Molecular and Cellular Biology, 2005
Members of the CELF family of RNA binding proteins have been implicated in alternative splicing regulation in developing heart. Transgenic mice that express a nuclear dominant-negative CELF protein specifically in the heart (MHC-CELF⌬) develop cardiac hypertrophy and dilated cardiomyopathy with defects in alternative splicing beginning as early as 3 weeks after birth. MHC-CELF⌬ mice exhibit extensive cardiac fibrosis, severe cardiac dysfunction, and premature death. Interestingly, the penetrance of the phenotype is greater in females than in males despite similar levels of dominant-negative expression, suggesting that there is sexspecific modulation of splicing activity. The cardiac defects in MHC-CELF⌬ mice are directly attributable to reduced levels of CELF activity, as crossing these mice with mice overexpressing CUG-BP1, a wild-type CELF protein, rescues defects in alternative splicing, the severity and incidence of cardiac hypertrophy, and survival. We conclude that CELF protein activity is required for normal alternative splicing in the heart in vivo and that normal CELF-mediated alternative splicing regulation is in turn required for normal cardiac function.
Nucleic Acids Research, 1991
We report here the isolation of the 5' end and the promoter region of the gene for chicken cartilage link protein, and demonstrate extensive heterogeneity of the leader sequence arising from differential utilization of multiple splice sites within the 5'-most exon. The 500-base pairs (bp) exon 1 consists of solely untranslated sequence and is followed by an intron >33 kilobase pairs (kb). Together, the five exons predict a gene size longer than 100 kb. Multiple transcription initiation sites were mapped 34, 46, 56, 66 and 76 bp downstream of a TATA-like motif. Sequence analysis revealed that in addition to the nonspliced variant, multiple mRNA species were generated by alternative splicing resulting in the exclusion of 92, 166, 170, 174 and 263 nucleotides (nt), respectively, from exon 1. Polymerase chain reaction confirmed the existence of various splice forms, and showed cell type-and developmental stage-specific expression for one group of them. Secondary structure predictions indicated that the leaders of the splice forms could form stable hairpin structures with different free energies of formation (up to AG = -1 10 kcal/mol), suggesting translational control. The splice variant detected in the largest amount had the least stable predicted hairpin (AG = -31.7 kcal/mol).
Proceedings of the National Academy of Sciences, 2015
Studies in several cell types have highlighted dramatic and diverse changes in mRNA processing that occur upon cellular stimulation. However, the mechanisms and pathways that lead to regulated changes in mRNA processing remain poorly understood. Here we demonstrate that expression of the splicing factor CELF2 (CUGBP, Elav-like family member 2) is regulated in response to T-cell signaling through combined increases in transcription and mRNA stability. Transcriptional induction occurs within 6 h of stimulation and is dependent on activation of NF-κB. Subsequently, there is an increase in the stability of the CELF2 mRNA that correlates with a change in CELF2 3′UTR length and contributes to the total signalinduced enhancement of CELF2 expression. Importantly, we uncover dozens of splicing events in cultured T cells whose changes upon stimulation are dependent on CELF2 expression, and provide evidence that CELF2 controls a similar proportion of splicing events during human thymic T-cell development. Taken together, these findings expand the physiologic impact of CELF2 beyond that previously documented in developing neuronal and muscle cells to T-cell development and function, identify unappreciated instances of alternative splicing in the human thymus, and uncover novel mechanisms for CELF2 regulation that may broadly impact CELF2 expression across diverse cell types.
Mechanisms of Development, 2001
Chicken embryonic stem (CES) cells are pluripotent cells derived from chicken early blastoderm. In order to identify new genes speci®cally expressed in these pluripotent cells, we have used a gene trap strategy and cloned a novel gene family called cENS for chicken Embryonic Normal Stem cell gene. The cENS genes expression decreases after induction of CES cells differentiation in culture and is restricted in vivo to the very early embryo. We have characterized three different cENS genes. One, cENS-1, is composed of an open reading frame inserted between two terminal direct repeats which are the common point of the cENS genes. cENS-1 encodes a protein identical to cERNI, a recently described protein. cENS-2 is a truncated form of cENS-1. cENS-3 presents two adjacent open reading frames coding respectively for env and pol related proteins. The presence of conserved direct repeats, of retrovirus related genes and the absence of introns argue in favor of a retroviral origin of the cENS genes. In the cENS we identi®ed a promoter region whose activity is strong in CES cells and decreases after induced differentiation showing a highly speci®c transcriptional activity speci®c of undifferentiated chicken embryonic stem cells. q
The neurofibromatosis type I pre-mRNA is a novel target of CELF protein-mediated splicing regulation
Nucleic Acids …, 2010
The CUG-BP and ETR-3 like factors (CELF) are a family of six highly conserved RNA-binding proteins that preferentially bind to UG-rich sequences. One of the key functions of these proteins is to mediate alternative splicing in a number of tissues, including brain, heart and muscle. To fully understand the function of CELF proteins, it is important to identify downstream targets of CELF proteins. In this communication, we report that neurofibromatosis type I (NF1) exon 23a is a novel target of CELF protein-mediated splicing regulation in neuron-like cells. NF1 regulates Ras signaling, and the isoform that excludes exon 23a shows 10 times greater ability to down-regulate Ras signaling than the isoform that includes exon 23a. Five of the six CELF proteins strongly suppress the inclusion of NF1 exon 23a. Over-expression or siRNA knockdown of these proteins in cell transfection experiments altered the levels of NF1 exon 23a inclusion. In vitro binding and splicing analyses demonstrate that CELF proteins block splicing through interfering with binding of U2AF65. These studies, combined with our previous investigations demonstrating a role for Hu proteins and TIA-1/TIAR in controlling NF1 exon 23a inclusion, highlight the complex nature of regulation of this important alternative splicing event.