Structure, mapping and expression of the human gene encoding the homeodomain protein, SIX2

Vertebrate genes with sequence similarity to the Drosophila homeobox gene, sine oculis (so), constitute the SIX family. There is notable expression of members of this family in anterior neural structures, and several SIX genes have been shown to play roles in vertebrate and insect development, or have been implicated in maintenance of the differentiated state of tissues. Mutations in three of these genes in man (SIX5, SIX6 and SIX3) are associated with severe phenotypes, and therefore, the cloning of other human genes from this family is of interest. We have cloned and characterised the gene that encodes human SIX2, elucidated its gene structure and conducted expression studies in a range of tissues. SIX2 is widely expressed in the late first-trimester fetus, but has a limited range of expression sites in the adult. The expression pattern of SIX2 and its localisation to chromosome 2p15-p16 will be of use in assessing its candidacy in human developmental disorders.     

Mesodermal expression of integrin α5β1 regulates neural crest development and cardiovascular morphogenesis

Integrin α5-null embryos die in mid-gestation from severe defects in cardiovascular morphogenesis, which stem from defective development of the neural crest, heart and vasculature. To investigate the role of integrin α5β1 in cardiovascular development, we used the Mesp1^Cre knock-in strain of mice to ablate integrin α5 in the anterior mesoderm, which gives rise to all of the cardiac and many of the vascular and muscle lineages in the anterior portion of the embryo. Surprisingly, we found that mutant embryos displayed numerous defects related to the abnormal development of the neural crest such as cleft palate, ventricular septal defect, abnormal development of hypoglossal nerves, and defective remodeling of the aortic arch arteries. We found that defects in arch artery remodeling stem from the role of mesodermal integrin α5β1 in neural crest proliferation and differentiation into vascular smooth muscle cells, while proliferation of pharyngeal mesoderm and differentiation of mesodermal derivatives into vascular smooth muscle cells was not defective. Taken together our studies demonstrate a requisite role for mesodermal integrin α5β1 in signaling between the mesoderm and the neural crest, thereby regulating neural crest-dependent morphogenesis of essential embryonic structures.     

The BON/CPN gene family represses cell death and promotes cell growth in Arabidopsis

Copines are calcium-dependent membrane-binding proteins that are highly conserved among protozoa, plants, nematodes and mammals. Although they are implicated in membrane trafficking and signal transduction, the functions of these proteins are not well understood. The Arabidopsis copine gene BON1/CPN1 was previously shown to negatively regulate a disease resistance (R) gene SNC1. Here we report that in Arabidopsis, as in other organisms, there is a family of copine genes, BON1, 2 and 3. Using double and triple mutant combinations we show that these three copine genes have overlapping functions essential for the viability of plants. The loss of function of BON1 combined with that of BON2 or BON3 leads to extensive cell death phenotypes resembling the hypersensitive response (HR) in defense responses. The resulting lethality can be suppressed by mutations in PAD4 or EDS1 which are required for R gene signaling and cell death control. Accession-dependent phenotypes of the mutant combinations suggest that the BON/CPN genes may together repress several R genes other than SNC1. Moreover, the mutant combinations exhibit developmental defects when R-gene-mediated defense responses are largely suppressed in pad4 and eds1 mutants. Thus, the copine family in Arabidopsis may have effects in promoting growth and development in addition to repressing cell death, and these two processes might be intricately intertwined.     

Muscle-specific loss of apoptosis-inducing factor leads to mitochondrial dysfunction, skeletal muscle atrophy, and dilated cardiomyopathy

Cardiac and skeletal muscle critically depend on mitochondrial energy metabolism for their normal function. Recently, we showed that apoptosis-inducing factor (AIF), a mitochondrial protein implicated in programmed cell death, plays a role in mitochondrial respiration. However, the in vivo consequences of AIF-regulated mitochondrial respiration resulting from a loss-of-function mutation in Aif are not known. Here, we report tissue-specific deletion of Aif in the mouse. Mice in which Aif has been inactivated specifically in cardiac and skeletal muscle exhibit impaired activity and protein expression of respiratory chain complex I. Mutant animals develop severe dilated cardiomyopathy, heart failure, and skeletal muscle atrophy accompanied by lactic acidemia consistent with defects in the mitochondrial respiratory chain. Isolated hearts from mutant animals exhibit poor contractile performance in response to a respiratory chain-dependent energy substrate, but not in response to glucose, supporting the notion that impaired heart function in mutant animals results from defective mitochondrial energy metabolism. These data provide genetic proof that the previously defined cell death promoter AIF has a second essential function in mitochondrial respiration and aerobic energy metabolism required for normal heart function and skeletal muscle homeostasis.     

The effector SIX8 is required for virulence of Fusarium oxysporum f.sp. cubense tropical race 4 to Cavendish banana

Plant pathogens employ effectors as molecular weapons to manipulate host immunity and facilitate colonization. Fusarium oxysporum f. sp. cubense is the agent of wilt disease in banana plantlets and four races of the pathogen have been identified based on the cultivar specificity. A total of 9 SIX genes have been detected in the genome of Foc TR4 and 6 genes detected in Foc1. Among these SIX genes, SIX2 and SIX8 are only detected in Foc TR4, not identified in Foc1. Expression profiles analysis revealed that SIX genes of Foc TR4 are highly induced after inoculation to Cavendish banana plantlets. Virulence analysis of the SIX2 and SIX8 knock-out mutants showed that SIX8 is required for the virulence of Foc TR4 while SIX2 has no obvious functions. Over expression of SIX8-FLAG proteins in the SIX8 knock-out mutant partly restored the virulence. Western blot analysis suggested that SIX8 could be secreted into the extracellular space and a signal peptide resided the N-terminal polypeptide sequence. This study provides some clues for further research on mechanism of SIX8 in regulating virulence of Foc TR4.     

Six class homeobox genes in drosophila belong to three distinct families and are involved in head development.

The vertebrate Six genes are homologues of the Drosophila homeobox gene sine oculis (so), which is essential for development of the entire visual system. Here we describe two new Six genes in Drosophila, D-Six3 and D-Six4, which encode proteins with strongest similarity to vertebrate Six3 and Six4, respectively. In addition, we report the partial sequences of 12 Six gene homologues from several lower vertebrates and show that the class of Six proteins can be subdivided into three major families, each including one Drosophila member. Similar to so, both D-Six3 and D-Six4 are initially expressed at the blastoderm stage in narrow regions of the prospective head and during later stages in specific groups of head midline neurectodermal cells. D-Six3 may also be essential for development of the clypeolabrum and several head sensory organs. Thus, the major function of the ancestral Six gene probably involved specification of neural structures in the cephalic region.     

Mice mutant in the DM domain gene Dmrt4 are viable and fertile but have polyovular follicles

Proteins containing the DM domain, a zinc finger-like DNA binding motif, have been implicated in sexual differentiation in diverse metazoan organisms. Of seven mammalian DM domain genes, only Dmrt1 and Dmrt2 have been functionally analyzed. Here, we report expression analysis and targeted disruption of Dmrt4 (also called DmrtA1) in the mouse. Dmrt4 is widely expressed during embryonic and postnatal development. However, we find that mice homozygous for a putative null mutation in Dmrt4 develop essentially normally, undergo full sexual differentiation in both sexes, and are fertile. We observed two potential mutant phenotypes in Dmrt4 mutant mice. First, ovaries of most mutant females have polyovular follicles, suggesting a role in folliculogenesis. Second, 25% of mutant males consistently exhibited copulatory behavior toward other males. We also tested potential redundancy between Dmrt4 and two other gonadally expressed DM domain genes, Dmrt1 and Dmrt7. We observed no enhancement of gonadal phenotypes in the double mutants, suggesting that these genes function independently in gonadal development.     

Genetic screens for genes controlling motor nerve-muscle development and interactions

Motor growth cones navigate long and complex trajectories to connect with their muscle targets. Experimental studies have shown that this guidance process critically depends on extrinsic cues. In the zebrafish embryo, a subset of mesodermal cells, the adaxial cells, delineates the prospective path of pioneering motor growth cones. Genetic ablation of adaxial cells causes profound pathfinding defects, suggesting the existence of adaxial cell derived guidance factors. Intriguingly, adaxial cells are themselves migratory, and as growth cones approach they migrate away from the prospective axonal path to the lateral surface of the myotome, where they develop into slow-twitching muscle fibers. Genetic screens in embryos stained with an antibody cocktail identified mutants with specific defects in differentiation and migration of adaxial cells/slow muscle fibers, as well as mutants with specific defects in axonal pathfinding, including exit from the spinal cord and pathway selection. Together, the genes underlying these mutant phenotypes define pathways essential for nerve and muscle development and interactions between these two cell types.     

Anterior organization of the Caenorhabditis elegans embryo by the labial-like Hox gene ceh-13

The Caenorhabditis elegans lin-39, mab-5 and egl-5 Hox genes specify cell fates along the anterior-posterior body axis of the nematode during postembryonic development, but little is known about Hox gene functions during embryogenesis. Here, we show that the C. elegans labial-like gene ceh-13 is expressed in cells of many different tissues and lineages and that the rostral boundary of its expression domain is anterior to those of the other Hox genes. By transposon-mediated mutagenesis, we isolated a zygotic recessive ceh-13 loss-of-function allele, sw1, that exhibits an embryonic sublethal phenotype. Lineage analyses and immunostainings revealed defects in the organization of the anterior lateral epidermis and anterior body wall muscle cells. The epidermal and mesodermal identity of these cells, however, is correctly specified. ceh-13(sw1) mutant embryos also show fusion and adhesion defects in ectodermal cells. This suggests that ceh-13 plays a role in the anterior organization of the C. elegans embryo and is involved in the regulation of cell affinities.