Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.23 (beta-galactosidase)
 14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The molecular determinants that direct gene expression to the ventricles of the heart are for the most part unknown. Additionally, little data is available on how the anterior/posterior axis of the heart tube is determined and whether the left and right atrial and ventricular chambers are assigned as part of this process. Utilizing myosin light chain-2 ventricular promoter/beta-galactosidase reporter transgenes, we have determined the minimal cis-acting sequences required for ventricular-specific gene expression. In multiple independent transgenic mouse lines, we found that both a 250 base pair myosin light chain-2 ventricular promoter fragment, as well as a dimerized 28 bp sub-element (HF-1) containing binding sites for HF1a and HF1b/MEF2 factors, directed ventricular-specific reporter expression from as early as the endogenous gene, at day 7.5-8.0 post coitum. While the endogenous gene is expressed uniformly throughout both ventricles, the transgenes were expressed in a right ventricular/conotruncal dominant fashion, suggesting that they contain only a subset of the elements which respond to positional information in the developing heart tube. Expression of the transgene was cell autonomous and its temporospatial characteristics not affected by mouse strain/methylation state of the genome. To determine whether ventricular-specific expression of the transgene was dependent upon regulatory genes required for correct ventricular differentiation, the 250 base pair transgene was bred into both retinoid X receptoralpha and Nkx2-5 null backgrounds. The transgene was expressed in both mutant backgrounds, despite the absence of endogenous myosin light chain-2 ventricular transcript in Nkx2-5 null embryos. Ventricular specification, as judged by transgene expression, appeared to occur normally in both mutants. Thus, the HF-1 element, directs chamber-specific transcription of a transgene reporter independently of retinoid X receptoralpha and Nkx2-5, and defines a minimal combinatorial pathway for ventricular chamber gene expression. The patterned expression of this transgene may provide a model system in which to investigate the cues that dictate anterior-posterior (right ventricle/left ventricle) gradients during mammalian heart development.
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PMID:An HF-1a/HF-1b/MEF-2 combinatorial element confers cardiac ventricular specificity and established an anterior-posterior gradient of expression. 867 19

Adenoviruses are very attractive vectors for gene transfer into the cardiac muscle; however, their promiscuous tissue tropism, leading to an ectopic expression of the transgene, is a considerable practical limitation. To restrict expression of a reporter gene in cultured cardiomyocytes and in the heart of the rat, we have constructed a recombinant adenovirus (Ad-MLC2 beta gal) containing the beta-galactosidase gene under the control of the rat ventricle-specific cardiac myosin light chain 2 (MLC-2v) promoter. We show in this work that the MLC-2v promoter inside the adenoviral genome retains its cardiac specificity in vitro in cultured cardiomyocytes as well as in vivo in the animal heart. Northern blot studies after Ad-MLC2 beta gal infection show significant transcription only in cells derived from the cardiac muscle and not from the skeletal muscle. Quantitative analysis of the beta-galactosidase activity in a number of cell lines also confirms this result. The level of beta-galactosidase expression in rat neonatal cardiomyocytes infected with Ad-MLC2 beta gal is 8% of that found when primary cells are infected with Ad-RSV beta gal (containing a beta-galactosidase gene under the control of the Rous sarcoma virus promoter). The cardiomyocytes-specific expression is also found after injection of Ad-MLC2 beta gal directly into the rat myocardium, although the viral genome can be detected by polymerase chain reaction (PCR) in other tissues. Lack of expression after direct injection into liver and skeletal muscle confirms these results. The use of a tissue-specific promoter is a first step to restrict transgene expression to a particular cell type of the targeted tissue.
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PMID:Expression from cardiomyocyte-specific promoter after adenovirus-mediated gene transfer in vitro and in vivo. 918 Nov 18

Pluripotent embryonic stem (ES) cells spontaneously differentiate via embryo-like aggregates into cardiomyocytes of pacemaker-, atrium- and ventricle-like type, which can be distinguished by their specific patterns of action potentials. It has been shown that retinoic acid (RA) treatment during ES cell differentiation increases the number of cardiomyocytes in a time- and concentration-dependent manner. In order to test the effect of RA on cardiomyocyte differentiation and specialization into ventricle-like cardiomyocytes, we studied gene expression of beta-galactosidase driven by the ventricular myosin light chain-2 (MLC-2v) promoter as an indicator for ventricular differentiation. Clones containing the stably integrated expression vector pGNA/MLC-2.1 were selected, which revealed an increase of beta-galactosidase activity in cardiomyocytes of embryoid bodies at day 7 + 16. RA, both, in the all-trans and in the 9-cis configuration resulted in a significant acceleration of cardiomyocyte differentiation and a transient increase of beta-galactosidase activity. To test whether this acceleration of cardiac differentiation and RA-induced increase of the MLC-2v promotor/beta-galactosidase activity reflects an increase of cardiac- and ventricle-specific gene expression, a semi-quantitative RT-PCR analysis was performed for alpha-cardiac myosin heavy chain (alpha-MHC) and MLC-2v genes. It was shown that both 10(-8) M and 10(-9) M RA resulted in an increased level of alpha-cardiac MHC and MLC-2v mRNA in embryoid bodies in early, but not in terminal developmental stages. This led us to the conclusion that the RA-induced accelerated expression of cardiac-specific genes results in an enhanced development of ventricular cardiomyocytes. An increased number of ventricle-like cells after RA treatment was also found by patch-clamp analysis. The number of cardiomyocytes with Purkinje- and ventricle-like properties was shown to be increased by RA, whereas the number of pacemaker- and atrium-like cells was reduced and early pacemaker cells were not quantitatively affected.
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PMID:Retinoic acid accelerates embryonic stem cell-derived cardiac differentiation and enhances development of ventricular cardiomyocytes. 922 Mar 39

The goal of this protocol is to describe a method for the dissection of mouse embryos and visualization of embryonic mouse ventricular chambers during heart development using ventricular specific fluorescent reporter knock-in mice (MLC-2v-tdTomato mice). Heart development involves a linear heart tube formation, the heart tube looping, and four chamber septation. These complex processes are highly conserved in all vertebrates. The mouse embryonic heart has been widely used for heart developmental studies. However, due to their extremely small size, dissecting mouse embryonic hearts is technically challenging. In addition, visualization of cardiac chamber formation often needs in situ hybridization, beta-galactosidase staining using LacZ reporter mice, or immunostaining of sectioned embryonic hearts. Here, we describe how to dissect mouse embryonic hearts and directly visualize ventricular chamber formation of MLC-2v-tdTomato mice using whole mount epifluorescent microscopy. With this method, it is possible to directly examine heart tube formation and looping, and four chamber formation without further experimental manipulation of mouse embryos. Although the MLC-2v-tdTomato reporter knock-in mouse line is used in this protocol as an example, this protocol can be applied to other heart-specific fluorescent reporter transgenic mouse lines.
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PMID:Analysis of Cardiac Chamber Development During Mouse Embryogenesis Using Whole Mount Epifluorescence. 3105 4