Drosophila GATA homologs

Pannier is related to GATA-1, a vertebrate transcription factor required for regulation of globin and other erythroid genes (Ramain, 1993 and Winick, 1993).

dGATAb (also known as Serpent), the other GATA homolog in Drosophila, is expressed is expressed in the fat body and is required for fat body gene expression. Serpent is also expressed in adult females in ovarian follicle cells. It regulates transcription of follicle specific proteins (Lossky, 1995). Serpent is required for hematopoiesis in Drosophila. serpent expression is seen in a patch of cells in the anterior portion of the mesodermal primordium. These cells invaginate with the portion of the ventral furrow that is anterior to the cephalic furrow. Slightly later these cells differentiate into prohemocytes and subsequently, become distributed throughout the body and differentiate as mature hemocytes. It is proposed that the mesodermal patch of srp constitutes the hemocyte primordium at blastoderm stage. These findings imply involvement of GATA in blood cell development and in endodermal differentiation. Such an involvement reflects an early phenomenon of metazoan evolution and may be common to most higher animals (Rehorn, 1996).

Pannier homologs in other insects

The Drosophila gene pannier (pnr) has been assigned to a new class of selector genes. It specifies pattern in the dorsal body. On the dorsal notum it is expressed in a broad medial domain and directly regulates transcription of the achaete-scute (ac-sc) genes driving their expression in small discrete clusters within this domain at the sites of each future bristle. This spatial resolution is achieved through modulation of Pnr activity by specific co-factors and by a number of discrete cis-regulatory enhancers in the ac-sc gene complex. Homologs of pnr and ac-sc have been isolated in Anopheles gambiae, a basal species of Diptera that diverged from Drosophila melanogaster (Dm) about 200 million years ago, and their expression patterns were examined. An ac-sc homolog of Anopheles, Ag-ASH, is expressed on the dorsal medial notum at the sites where sensory organs emerge in several domains that are identical to those of the pnr homolog, Ag-pnr. This suggests that activation of Ag-ASH by Ag-Pnr has been conserved. Indeed, when expressed in Drosophila, Ag-pnr is able to mimic the effects of ectopic expression of Dm-pnr and induce ectopic bristles. These results are discussed in the context of the gene duplication events and the acquisition of a modular promoter, that may have occurred at different times in the lineage leading to derived species such as Drosophila. The bristle pattern of Anopheles correlates in a novel fashion with the expression domains of Ag-pnr/Ag-ASH. While precursors for the sensory scales can arise anywhere within the expression domains, bristle precursors arise exclusively along the borders. This points to the existence of specific positional information along the borders, and suggests that Ag-pnr specifies pattern in the medial, dorsal notum, as in Drosophila, but via a different mechanism (Wülbeck, 2002).

Ag-ASH appears closest to Drosophila l'sc. Sequence analysis has revealed that 81% of the amino acids in the bHLH domain are identical to those of the Drosophila l'sc protein. Outside of this functional domain, amino acid sequence conservation is low (ranging from 20%-27% for the amino (N)-terminal portion to 25%-38% for the carboxy (C)-terminal part). A single stretch of 15 conserved amino acids, which appears to be restricted to insect ac-sc proteins, can be seen at the C terminus. The central tyrosine of this sequence has changed in the butterfly Precis coenia (Wülbeck, 2002).

The screening procedure used allowed the isolation of a single Anopheles ASC homolog, Ag-ASH, but examination of the recently published genome of this species reveals the existence of an asense gene. Ag-ASH is closest to Drosophila l'sc, but may be representative of an ancestral gene, which was present prior to the duplication events that gave rise to l'sc, sc and ac. This may have taken place after separation of the Nematocera (including the mosquitoes) and Brachycera (including Drosophila and Ceratitis), two lineages that diverged about 200 million years ago. A single ASC homolog has been described in the butterfly Precis coenia. When expressed in Drosophila, Ag-ASH has a conserved and strong, proneural function (Wülbeck, 2002).

The pnr gene of Drosophila comprises two zinc fingers characteristic of the GATA family of transcription proteins, and a C-terminal domain bearing two alpha helices. The protein contains two zinc fingers that are very strongly conserved. The proteins are, however, quite divergent in the C-terminal domain. The proteins of Ceratitis and Anopheles carry a single alpha helix, in contrast to the two in Drosophila (Wülbeck, 2002).

In Drosophila, pnr is expressed in a conserved broad medial domain but activates ac and sc in discrete proneural clusters within this domain. The ac-sc genes of Drosophila are organized into a complex containing multiple enhancer regions, each of which independently regulates expression in one or a small number of proneural clusters. In this species three proneural clusters arise in the domain of pnr expression and Pnr has been shown to directly activate ac-sc in the dorso-central cluster, through binding to a cis-regulatory sequence just upstream of ac. It is not entirely understood how the broad domain of Pnr is translated into the small clusters of ac-sc expression, but this is at least in part achieved through interaction of Pnr with regulatory co-factors. The spatially complex expression of sc in Calliphora and Ceratitis suggests that the ASC genes of these species may also have modular promoters. Furthermore, the expression domain of pnr in these species is conserved with that of Drosophila (Wülbeck, 2002).

In contrast, the regulatory interactions between the two genes appear to have diverged in Anopheles since Ag-ASH is expressed in all Ag-pnr-expressing cells. The common domains of expression suggest that Ag-Pnr may activate Ag-ASH in every cell in which it is expressed, in a simple straightforward fashion. This observation raises two possibilities. (1)for the regulation of Ag-ASH, Ag-Pnr may not associate with the various co-factors known to modulate its activity in Drosophila; (2) in order to be activated in all Ag-pnr-expressing cells, Ag-ASH would not need to have a modular promoter structure like that of the Drosophila locus, and could have a less complex organization. If so, the acquisition of position-specific enhancers may have occurred after the separation of Nematocera and Brachycera, at a time when further gene duplication events appear to have taken place. In addition, modulation of Pnr activity through the use of different co-factors may have accompanied the acquisition of cis-regulatory enhancer sequences in the lineage leading to Drosophila (Wülbeck, 2002).

Despite the inferred simple regulatory interaction between Ag-Pnr and Ag-ASH, it is remarkable that the effects of mis-expression of Ag-pnr in Drosophila are almost identical to those caused by mis-expression of Dm-pnr. For example, ectopic expression of either Dm-pnr or Ag-pnr on the lateral notum, causes the development of a tuft of ectopic dorso-central bristles. This is due to an expansion of the activity of the dorso-central enhancer element known to be regulated by Dm-Pnr. This result suggests that Ag-Pnr is able to recognise the relevant regulatory modules of the Drosophila ASC promoter; this may indicate that these enhancers are derived from an ancestral regulatory sequence also present in Anopheles. Alternatively, a number of regulatory modules may in fact be present in the Anopheles promoter and govern expression in the various domains on the notum. Further understanding of the structure and regulation of Ag-ASH will require investigation of regulatory sequences from this organism. The ectopic expression assay also indicates that Ag-Pnr is probably able to associate with Drosophila co-factors such as U-shaped and Chip. It has been shown that the N-terminal zinc finger of Dm-Pnr associates with U-shaped, while two C-terminal helical structures are components mediating association with Chip. The two zinc fingers are strongly conserved in Ag-Pnr, and there is a single alpha helix. Thus Ag-Pnr appears to contain the relevant binding regions for these two factors. This complexity of the Ag-pnr protein may indicate association with endogenous co-factors, perhaps in a different tissue (Wülbeck, 2002).

In Drosophila, it has been demonstrated, that pnr and the iro-C genes are selector genes involved in the subdivision of the dorsal component of segments of the head, thorax and abdomen of the adult into medial and lateral domains. While pnr regulates the pattern of the medial domain of the dorsal mesonotum, patterning of the lateral half is regulated by the iro-C genes. Thus, when either Dm-pnr or Ag-pnr is expressed from an early stage in the entire notum of Drosophila, only structures corresponding to the medial notum are formed: the lateral region fails to develop. Ubiquitous expression specifies a single medial domain thought to include cells originally destined to form the lateral region. In addition Ag-pnr is expressed in the medial, but not the lateral, mesonotum of Anopheles, consistent with a conserved function in the medial domain. Thus the selector gene function of pnr may have been conserved. The function of proteins of other selector genes of Anopheles, such as engrailed, has been shown to be conserved (Wülbeck, 2002).

The precursors of the sensory scales on the notum of Anopheles are distributed in a random fashion within the domains of expression of Ag-pnr/Ag-ASH. In some respects the sensory scales resemble the small bristles or microchaetes of cyclorraphous Diptera, which are often randomly distributed although sometimes lined up into rows. However, in the latter species they arise later than the large bristles or macrochaetes, from a second period of ac-sc expression, and are consequently positioned closer to one another than are the macrochaetes. In contrast, the precursors of scales and bristles appear to arise simultaneously in Anopheles, which is consistent with the fact that they are equidistant from each other in the imago. In cyclorraphous flies, the macrochaete pattern is the result of spatially complex sc (ac) expression: one (or a very small number) of bristle(s) develops from each small cluster (or stripe) of sc (ac) expression. In Anopheles, however, the patterning mechanism is different: remarkably, the precursors of the bristles are exclusively positioned along the borders of the expression domains. Thus the positions of the rows of AC and DC bristles, as well as the PST and SC bristles, are coincident with the borders of the four domains of Ag-pnr/Ag-ASH expression. This suggests that the boundaries of Ag-ASH/Ag-pnr expression convey specific positional information causing neural precursors to develop into bristles rather than sensory scales (Wülbeck, 2002).

Two observations in Drosophila may be relevant to this phenomenon. (1) Some of the macrochaete precursors arise from the edges of the corresponding proneural clusters of ac-sc expression, an observation that has been linked to distance from the source of the signaling molecules Wingless and Decapentaplegic. The expression pattern of these molecules in Anopheles is not yet known. (2) It has been demonstrated that the border between pnr-expressing and non-expressing cells does in fact display special properties. Cells of the medial domain manifest unique adhesive characteristics that prevent them from mixing with cells of the lateral domain. So, as for compartment boundaries, this interface between cells expressing pnr and those expressing iro may be an important patterning boundary. It has indeed been shown to be required for the growth and patterning of the Drosophila eye. Interestingly, the five macrochaetes on the medial notum of Drosophila are pnr-dependent, and they are all positioned on the lateral border of the domain of pnr expression. Experimentally contrived expression of ac-sc inside the pnr domain, however, results in the formation of ectopic macrochaetes, indicating that macrochaete formation in Drosophila, is not dependent on special properties at the border. Furthermore the prescutellar bristle of Ceratitis and the AC row of bristles in Calliphora, arise from sc-expressing cells situated inside the pnr expression domain (Wülbeck, 2002).

Although the bristles on the notum of Anopheles are aligned into rows, the number and position of bristles within a row varies greatly between individuals, a feature that is thought to be ancestral. In contrast, species of cyclorraphous Schizophora have very defined rows in which the number and position of bristles varies little if at all. The stereotyped notal bristle patterns of species such as Drosophila are thought to be derived from an ancestral pattern of four longitudinal rows of bristles, still present in many extant species of Schizophora. These include the AC and DC bristle rows that are in the medial domain of the notum. So, for example, the two DC bristles of Drosophila would be vestiges of the DC row. Whether the rows of bristles seen in some families of Nematocera such as the Culicidae, are in any way related by ancestry to the four rows of Schizophoran flies, is more difficult to assess. Nevertheless the DC row of Anopheles is positioned on the lateral border of the Ag-pnr expression domain, as in Ceratitis, Calliphora and Drosophila, which may indicate a common origin for this row. If so, this would mean that an ancestral pattern of bristle rows was already present in a common ancestor of the Brachycera and at least some families of Nematocera (Wülbeck, 2002).

These results indicate a conserved function for pnr in the regulation of the bristle pattern on the medial notum. This argues in favour of an ancient role for pnr as a selector gene specifying the dorsal medial pattern. The nature of the regulatory interactions between Pnr and its target genes ac-sc appears to have changed, however, over evolutionary time. It is hypothesized that in Culicid mosquitoes, which have fewer ac-sc genes, the regulatory regions of this locus may not be organized in a modular fashion. Evolution of the stereotyped bristle patterns characteristic of species such as Drosophila and Ceratitis may have entailed the acquisition of a number of additional factors. These would include gene duplication within the ASC and the co-option of cis-regulatory sequences. Co-factors for Pnr, such as Ush and Chip, are also likely to have been co-opted for use in constructing the notal pattern at a later evolutionary stage, although the current results suggest that Ag-Pnr has the requisite domains for association with these proteins. In the lineage leading to Drosophila, these different levels of regulation might have been superimposed onto an ancestral patterning mechanism, similar to that of Anopheles, at different times in the 200 million years separating Drosophila from the Nematocera (Wülbeck, 2002).

C. elegans Pannier homologs

Epidermal cells are generated during C. elegans embryogenesis by several distinct lineage patterns. These patterns are controlled by maternal genes that determine the identities of early embryonic blastomeres. The embryonically expressed gene elt-1 encodes a GATA-like transcription factor. ELT-1 protein contains two zinc finger domains and is 60% identical in these domains to vertebrate GATA factors, such as GATA-1, 2 and 3 from the mouse. elt-1 mutants lack hypodermal cells and the gene is required for the production of epidermal cells by each of four independent hypodermal lineages. Depending on their lineage history, cells that become epidermal in wild-type embryos become either neurons or muscle cells in elt-1 mutant embryos. The ELT-1 protein is expressed in epidermal cells and in their precursors. It is proposed that elt-1 functions at an early step in the specification of epidermal cell fates. It is intriguing that the elt-1 gene and the pannier gene of Drosophila encode similar GATA-like transcription factors. Severe mutations in pannier cause embryonic lethality, and the arrested embryos appear to lack epidermal cells. It is thus possible that pannier and elt-1 have evolutionarily related functions in epidermal tissue specification (Page, 1997).

A gene encoding elt-3, a new member of the Caenorhabditis elegans GATA transcription factor family, has been identifed. The predicted ELT-3 polypeptide contains a single GATA-type zinc finger (C-X2-C-X17-C-X2-C) along with a conserved adjacent basic region. elt-3 is slightly more closely related to pannier, expressed in Drosophila ectoderm, than to Drosophila serpent, but the significance of this small difference is questionable. elt-3 mRNA is present in all stages of C. elegans development but is most abundant in embryos. Reporter gene analysis and antibody staining show that elt-3 is first expressed in the dorsal and ventral hypodermal cells, and in hypodermal cells of the head and tail, immediately after the final embryonic cell division that gives rise to these cells. No expression is seen in the lateral hypodermal (seam) cells. elt-3 expression is maintained at a constant level in the epidermis until the 2(1/2)-fold stage of development, after which reporter gene expression declines to a low level and endogenous protein can no longer be detected by specific antibody. A second phase of elt-3 expression in cells immediately anterior and posterior to the gut begins in pretzel-stage embryos. elt-1, coding for another GATA factor, and lin-26, a multiple zinc finger transcription factor, are two genes known to be important in specification and maintenance of hypodermal cell fates. elt-1 is required for the formation of most, but not all, elt-3-expressing cells. In contrast, lin-26 function does not appear necessary for elt-3 expression. The candidate homolog of elt-3 has been characterized in the nematode Caenorhabditis briggsae. Many features of the elt-3 genomic and transcript structure are conserved between the two species, suggesting that elt-3 is likely to perform an evolutionarily significant function during development (Gilleard, 1999).

In vertebrates, transcriptional regulators of the GATA family appear to have a conserved function in differentiation and organ development. GATA-1, -2 and -3 are required for different aspects of hematopoiesis, while GATA-4, -5 and -6 are expressed in various organs of endodermal origin, such as intestine and liver, and are implicated in endodermal differentiation. GATA-1 is required for primitive and definitive erythropoiesis, GATA-2 for early hematopoiesis, and GATA-3 is implicated in the differentiation of T-lymphocytes. Pannier and Serpent are recently diverged on an evolutionary scale, each similarly related in sequence to all vertebrate GATA proteins (Rehorn, 1996 and references).

Vertebrate pannier-related genes

Vertebrate GATA proteins are extensively dealt with at the Serpent site.

The GATA family members fall into two subgroups: GATA-1, -2 and -3 are contained in one, represented in Drosophila by Pannier, and GATA-4, -5 and -6 in the other, represented by Serpent (but see Rehorn, 1996 above). These subgroups are defined in terms of both their sequence homology and expression patterns: GATA-1, -2 and -3 predominate in blood and ectodermal derivatives and GATA-4, -5 and -6 predominate in heart and endodermal tissues. GATA-2 is found in endothelial cells, trophoblasts, and the nervous system, as well as in hematopoietic stem cells, mast cells, and primitive erythroblasts. GATA-2-deficient ES cells do not give rise to any cells of the hematopoietic lineage in adult chimeras, thus showing that GATA-2 plays a critical role in the maintenance and proliferation of hematopoietic stem cells. Xenopus GATA-2 is present in Xenopus oocytes as a nuclear complex that is maintained throughout early development. It is stored in oocytes and embryos as a non-chromatin-bound complex. However, some oocyte GATA-2 is functional as a transcription factor because it can function on a chicken hemoglobin promoter when injected into oocytes. It is possible that complexing of the maternal Xenopus GATA-2 plays a role in development by restricting the range of GATA sequences to which it can bind, thereby limiting activation only to those genes with very high affinity sites (Partington, 1997).

The mechanisms regulating vertebrate heart and endoderm development have recently become the focus of intense study. Evidence is presented from both loss- and gain-of-function experiments that the zinc finger transcription factor Gata5 is an essential regulator of multiple aspects of heart and endoderm development. Zebrafish Gata5 is encoded by the faust locus. Analysis of faust mutants indicates that early in embryogenesis Gata5 is required for the production of normal numbers of developing myocardial precursors, as well as the expression of normal levels of several myocardial genes including nkx2.5. Later, Gata5 is necessary for the elaboration of ventricular tissue. Gata5 is required for the migration of the cardiac primordia to the embryonic midline and for endodermal morphogenesis. Significantly, overexpression of gata5 induces the ectopic expression of several myocardial genes including nkx2.5 and can produce ectopic foci of beating myocardial tissue. Together, these results implicate zebrafish Gata5 in controlling the growth, morphogenesis, and differentiation of the heart and endoderm and indicate that Gata5 regulates the expression of the early myocardial gene nkx2.5 (Reiter, 1999).

Gata5 is capable of activating transcription from a wide range of myocardial promoters; gata5 mutants display marked defects in the expression of many genes encoding components of the myocardial sarcomere (e.g., cmlc1, cmlc2, vmhc, cardiac troponin T, tropomyosin). However, it is not clear at this point which of these genes are direct targets of Gata5 and which require intermediate Gata5-dependent regulators. In fact, there may be no clear division between these cases as Gata5 may act both directly and indirectly on a single promoter. For example, Gata5 may participate in the induction of nkx2.5 and also bind cooperatively with Nkx2.5 to regulatory elements of a wide range of myocardial genes. It is also interesting to note the different requirements various sarcomeric protein genes have for Gata5. For example, although the expression patterns of cmlc1 and cmlc2 are indistinguishable in wild-type embryos, cmlc1 expression is more severely reduced than cmlc2 expression in fau mutants. Perhaps these differences reflect different affinities of Gata5 for the respective cis regulatory elements (Reiter, 1999).

The zinc finger transcription factors GATA-2 and GATA-3 are expressed in trophoblast giant cells. They regulate transcription from the mouse placental lactogen I gene promoter in a transfected trophoblast cell line. In trophoblast giant cells in vivo, both of these factors regulate transcription of the placental lactogen I gene, as well as the related proliferin gene. Placentas lacking GATA-3 accumulate placental lactogen I and proliferin mRNAs to a level 50% below that reached in the wild-type placenta. Mutation of the GATA-2 gene has a similar effect on placental lactogen I expression, but leads to a markedly greater reduction (5- to 6-fold) in proliferin gene expression. Placentas lacking GATA-2 secrete significantly less angiogenic activity than wild-type placentas, as measured in an endothelial cell migration assay, consistent with a reduction in expression of the angiogenic hormone proliferin. Furthermore, within the same uterus, the decidual tissue adjacent to mutant placentas displays markedly reduced neovascularization compared to the decidual tissue next to wild-type placentas. These results indicate that GATA-2 and GATA-3 are important in vivo regulators of trophoblast-specific gene expression and placental function, and reveal a difference in the effect of these two factors in regulating the synthesis of related placental hormones (Ma, 1997).

The ventralizing factor bone morphogenetic protein 4 (BMP-4) can inhibit Xenopus neurogenesis. The erythroid transcription factor GATA-1 functions downstream of the BMP-4 signaling pathway and mediates BMP-4-induced erythropoiesis. Similar to BMP-4, GATA-1b inhibits neuralization of Xenopus animal cap (AC) cells. The neural inhibition is not seen with GATA-1a, although both GATA-1a and GATA-1b RNAs are translated at the same efficiency and induce globin expression equally in AC cells. GATA-1b RNA injection into AC cells neither induces expression of Xbra (a general mesoderm marker) nor affects expression of XK81 (epidermal keratin) or BMP-4 and Xvent-1 (two ventral markers). These data suggest that GATA-1b retains the epidermal fate of the AC. Intact GATA-1b protein is required for both inhibition of neurogenesis and induction of globin expression. These findings indicate that GATA-1b can function in ectoderm to specifically regulate neural inducing mechanisms, apparently related to the expression of chordin, a neuralizing gene. Furthermore, tadpole stage embryos injected with GATA-1b are devoid of all dorsoanterior structures including neural tissue. This report provides evidence that the two transcription factors, derived from a recent genome duplication, share a common biological activity (stimulation of erythropoiesis) while also exhibiting a distinct function (inhibition of neurogenesis) (Xu, 1997).

The molecular mechanisms specifying patterns of gene expression in the vertebrate brain, which in turn determine the developmental fates of specific neurons, are yet to be clearly defined. Individual members of a recently identified family of transcriptional regulatory proteins, the GATA factors, are required for the differentiation of certain hematopoietic cell lineages. Two of the members of this gene family, GATA-2 and GATA-3, are expressed within discrete cell populations of the chicken optic tectum during embryogenesis; they have highly restricted patterns of expression in the developing chicken brain. The induction of GATA factor expression within specific cell layers parallels the well established spatial (rostral to caudal) and temporal pattern of optic tectum development. The observation that both the timing of appearance and the localization of expression of GATA-2 and GATA-3 are correlated with optic tectum development suggests that these transcription factors may be associated with the initiation of gene transcription required for the determination of specific neuronal fates within visual areas of the vertebrate brain (Kornhauser, 1994).

The tissue-specific transcription factor GATA-1 is a key regulator of red blood cell differentiation. One seemingly contradictory aspect of GATA-1 function is that while it is abundant in erythroid progenitor cells prior to the onset of overt differentiation, it does not significantly activate known GATA-1 target genes in those cells. In primary progenitor cells, GATA-1 protein is predominantly located in the cytoplasm, while induction of differentiation causes its rapid relocalization to the nucleus, suggesting that nuclear translocation constitutes an important regulatory step in GATA-1 activation. An ectopically expressed GATA-1/estrogen receptor fusion protein (GATA-1/ER) in red blood cell progenitors accelerates red blood cell differentiation, and concomitantly suppresses cell proliferation. Estrogen conditionally controls the nuclear translocation of GATA-1/ER, as well as its transcriptional activation. These phenotypic effects are accompanied by a simultaneous suppression of c-myb and GATA-2 transcription, two genes thought to be involved in the proliferative capacity of hematopoietic progenitor cells. Thus, GATA-1 appears to promote differentiation in committed erythroid progenitor cells both by inducing differentiation-specific genes and by simultaneously suppressing genes involved in cell proliferation (Briegel, 1996).

Co-occupancy by multiple cardiac transcription factors identifies transcriptional enhancers active in heart

Identification of genomic regions that control tissue-specific gene expression is currently problematic. ChIP and high-throughput sequencing (ChIP-seq) of enhancer-associated proteins such as p300 identifies some but not all enhancers active in a tissue. This study shows that co-occupancy of a chromatin region by multiple transcription factors (TFs) identifies a distinct set of enhancers. GATA-binding protein 4 (GATA4), NK2 transcription factor-related, locus 5 (NKX2-5), T-box 5 (TBX5), serum response factor (SRF), and myocyte-enhancer factor 2A (MEF2A), referred to as 'cardiac TFs,' have been hypothesized to collaborate to direct cardiac gene expression. Using a modified ChIP-seq procedure, chromatin occupancy by these TFs and p300 were defined genome wide and unbiased support for this hypothesis is provided. This principle was used to show that co-occupancy of a chromatin region by multiple TFs can be used to identify cardiac enhancers. Of 13 such regions tested in transient transgenic embryos, seven (54%) drove cardiac gene expression. Among these regions were three cardiac-specific enhancers of Gata4, Srf, and swItch/sucrose nonfermentable-related, matrix-associated, actin-dependent regulator of chromatin, subfamily d, member 3 (Smarcd3), an epigenetic regulator of cardiac gene expression. Multiple cardiac TFs and p300-bound regions were associated with cardiac-enriched genes and with functional annotations related to heart development. Importantly, the large majority (1,375/1,715) of loci bound by multiple cardiac TFs did not overlap loci bound by p300. These data identify thousands of prospective cardiac regulatory sequences and indicate that multiple TF co-occupancy of a genomic region identifies developmentally relevant enhancers that are largely distinct from p300-associated enhancers (He, 2011).

The cardiac transcription network modulated by Gata4, Mef2a, Nkx2.5, Srf, histone modifications, and microRNAs

The transcriptome, as the pool of all transcribed elements in a given cell, is regulated by the interaction between different molecular levels, involving epigenetic, transcriptional, and post-transcriptional mechanisms. However, many previous studies investigated each of these levels individually, and little is known about their interdependency. A systems biology study is presented integrating mRNA profiles with DNA-binding events of key cardiac transcription factors (Gata4, Mef2a, Nkx2.5, and Srf), activating histone modifications (H3ac, H4ac, H3K4me2, and H3K4me3), and microRNA profiles obtained in wild-type and RNAi-mediated knockdown. Finally, conclusions primarily obtained in cardiomyocyte cell culture were confirmed in a time-course of cardiac maturation in mouse around birth. Insights are provided into the combinatorial regulation by cardiac transcription factors and show that they can partially compensate each other's function. Genes regulated by multiple transcription factors are less likely differentially expressed in RNAi knockdown of one respective factor. In addition to the analysis of the individual transcription factors, it was found that histone 3 acetylation correlates with Srf- and Gata4-dependent gene expression and is complementarily reduced in cardiac Srf knockdown. Further, it was found that altered microRNA expression in Srf knockdown potentially explains up to 45% of indirect mRNA targets. Considering all three levels of regulation, an Srf-centered transcription network is presented providing on a single-gene level insights into the regulatory circuits establishing respective mRNA profiles. In summary, this study shows the combinatorial contribution of four DNA-binding transcription factors in regulating the cardiac transcriptome and provide evidence that histone modifications and microRNAs modulate their functional consequence. This opens a new perspective to understand heart development and the complexity cardiovascular disorders (Schlesinger, 2011; full text of article).

pannier: Biological Overview | Regulation | Developmental Biology | Effects of Mutation | References

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