Analysis of chromosomal localization of loci controlling milk production traits in cattle

Analysis of the pattern of the chromosomal localization of quantitative trait loci (QTLs) is necessary for comprehensively understanding their functions. The chromosomal localization of QTLs controlling milk production traits has been studied in cattle chromosomes. The distribution of QTLs between chromosomes has proved to be binomial. Their distribution along each chromosome was, in general, uniform, except for the QTLs controlling the somatic cell score (SCS), which tended towards telomeric location. However, there are chromosomes either enriched with or particularly poor in QTLs. The QTL distribution patters are the most similar for the milk yield (M) and milk protein yield (P) and for milk fat yield (F) and milk fat content (%F). The pattern of the SCS QTLs stands out among those of other QTLs. The distance between the QTLs of contrasting traits is the shortest for M and P QTLs, longer for M and milk protein content (%P) QTLs, and still longer for M and %F QTLs, which may be explained by QTL pleiotropy, a common phenomenon in cattle.     

LGI proteins in the nervous system

The development and function of the vertebrate nervous system depend on specific interactions between different cell types. Two examples of such interactions are synaptic transmission and myelination. LGI1-4 (leucine-rich glioma inactivated proteins) play important roles in these processes. They are secreted proteins consisting of an LRR (leucine-rich repeat) domain and a so-called epilepsy-associated or EPTP (epitempin) domain. Both domains are thought to function in protein–protein interactions. The first LGI gene to be identified, LGI1, was found at a chromosomal translocation breakpoint in a glioma cell line. It was subsequently found mutated in ADLTE (autosomal dominant lateral temporal (lobe) epilepsy) also referred to as ADPEAF (autosomal dominant partial epilepsy with auditory features). LGI1 protein appears to act at synapses and antibodies against LGI1 may cause the autoimmune disorder limbic encephalitis. A similar function in synaptic remodelling has been suggested for LGI2, which is mutated in canine Benign Familial Juvenile Epilepsy. LGI4 is required for proliferation of glia in the peripheral nervous system and binds to a neuronal receptor, ADAM22, to foster ensheathment and myelination of axons by Schwann cells. Thus, LGI proteins play crucial roles in nervous system development and function and their study is highly important, both to understand their biological functions and for their therapeutic potential. Here, we review our current knowledge about this important family of proteins, and the progress made towards understanding their functions.     

Identification of new protein-protein interactions involving the products of the chromosome- and plasmid-encoded type IV secretion loci of the phytopathogen Xanthomonas axonopodis pv. citri

The recently sequenced genome of the bacterial plant pathogen Xanthomonas axonopodis pv. citri contains two virB gene clusters, one on the chromosome and one on a 64-kb plasmid, each of which codes for a previously uncharacterized type IV secretion system (T4SS). Here we used a yeast two-hybrid assay to identify protein-protein interactions in these two systems. Our results revealed interactions between known T4SS components as well as previously uncharacterized interactions involving hypothetical proteins coded by open reading frames in the two X. axonopodis pv. citri virB loci. Our results indicate that both loci may code for previously unidentified VirB7 proteins, which we show interact with either VirB6 or VirB9 or with a hypothetical protein coded by the same locus. Furthermore, a set of previously uncharacterized Xanthomonas proteins have been found to interact with VirD4, whose gene is adjacent to the chromosomal virB locus. The gene for one member of this family is found within the chromosomal virB locus. All these uncharacterized proteins possess a conserved 120-amino-acid domain in their C termini and may represent a family of cofactors or substrates of the Xanthomonas T4SS.     

Mapping of Human DNA-binding Nuclear Protein (NP220) to Chromosome Band 2p13.1-p13.2 and Its Relation to Matrin 3

Human NP220 (hNP220) is a novel DNA-binding nuclear protein, which has an arginine/serine-rich motif and polypyrimidine tract-binding motif, and NP220s and matrin 3 are thought to form a novel family of nuclear proteins. We have determined a chromosomal localization of the cDNA encoding human NP220 to 2p13.1-p13.2 by using fluorescence in situ hybridization. Human matrin 3 cDNA was mapped to chromosomes 1p13.1-p21.1 and 5q31.3, demonstrating that these novel nuclear proteins with similar functions are on different chromosomes.     

Postzygotic isolation involves strong mitochondrial and sex-specific effects in Tigriopus californicus,a species lacking heteromorphic sex chromosomes

Detailed studies of the genetics of speciation have focused on a few model systems, particularly Drosophila. The copepod Tigriopus californicus offers an alternative that differs from standard animal models in that it lacks heteromorphic chromosomes (instead, sex determination is polygenic) and has reduced opportunities for sexual conflict, because females mate only once. Quantitative trait loci (QTL) mapping was conducted on reciprocal F₂ hybrids between two strongly differentiated populations, using a saturated linkage map spanning all 12 autosomes and the mitochondrion. By comparing sexes, a possible sex ratio distorter was found but no sex chromosomes. Although studies of standard models often find an excess of hybrid male sterility factors, we found no QTL for sterility and multiple QTL for hybrid viability (indicated by non-Mendelian adult ratios) and other characters. Viability problems were found to be stronger in males, but the usual explanations for weaker hybrid males (sex chromosomes, sensitivity of spermatogenesis, sexual selection) cannot fully account for these male viability problems. Instead, higher metabolic rates may amplify deleterious effects in males. Although many studies of standard speciation models find the strongest genetic incompatibilities to be nuclear–nuclear (specifically X chromosome–autosome), we found the strongest deleterious interaction in this system was mito–nuclear. Consistent with the snowball theory of incompatibility accumulation, we found that trigenic interactions in this highly divergent cross were substantially more frequent (>6 × ) than digenic interactions. This alternative system thus allows important comparisons to studies of the genetics of reproductive isolation in more standard model systems.     

In vivo assay for protein-protein interactions using Drosophila chromosomes

The ability of a chimeric HP1-Polycomb (Pc) protein to bind both to heterochromatin and to euchromatic sites of Pc protein binding was exploited to detect stable protein-protein interactions in vivo. Previously, we showed that endogenous Pc protein was recruited to ectopic heterochromatic binding sites by the chimeric protein. Here, we examine the association of other Pc group (Pc-G) proteins. We show that Posterior sex combs (Psc) protein also is recruited to heterochromatin by the chimeric protein, demonstrating that Psc protein participates in direct protein-protein interaction with Pc protein or Pc-associated protein. In flies carrying temperature-sensitive alleles of Enhancer of zeste[E(z)] the general decondensation of polytene chromosomes that occurs at the restrictive temperature is associated with loss of binding of endogenous Pc and chimeric HP1-Polycomb protein to euchromatin, but binding of HP1 and chimeric HP1-Polycomb protein to the heterochromatin is maintained. The E(z) mutation also results in the loss of chimera-dependent binding to heterochromatin by endogenous Pc and Psc proteins at the restrictive temperature, suggesting that interaction of these proteins is mediated by E(z) protein. A myc-tagged full-length Suppressor 2 of zeste [Su(z)2] protein interacts poorly or not at all with ectopic Pc-G complexes, but a truncated Su(z)2 protein is strongly recruited to all sites of chimeric protein binding. Trithorax protein is not recruited to the heterochromatin by the chimeric HP1-Polycomb protein, suggesting either that this protein does not interact directly with Pc-G complexes or that such interactions are regulated. Ectopic binding of chimeric chromosomal proteins provides a useful tool for distinguishing specific protein-protein interactions from specific protein-DNA interactions important for complex assembly in vivo.     

The program of sex chromosome pairing in meiosis is highly conserved across marsupial species: implications for sex chromosome evolution

Marsupials present a series of genetic and chromosomal features that are highly conserved in very distant species. One of these features is the absence of a homologous region between X and Y chromosomes. According to this genetic differentiation, sex chromosomes do not synapse during the first meiotic prophase in males, and a special structure, the dense plate, maintains sex chromosome association. In this report we present results on the process of meiotic sex chromosome pairing obtained from three different species, Thylamys elegans, Dromiciops gliroides, and Rhyncholestes raphanurus, representing the three orders of American marsupials. We have investigated the relationships between the axial structures organized along sex chromosomes and the formation of the dense plate. We found that in the three species the dense plate arises as a modification of sex chromosomal axial elements, but without the involvement of other meiotic axial structures, such as the cohesin axes. Considering the phylogenetic relationships among the marsupials studied here, our data reinforce the idea that the dense plate emerged early in marsupial evolution as an efficient mechanism to ensure the association of the nonhomologous sex chromosomes. This situation could have influenced the further evolution of sex chromosomes in marsupials.     

Analysis of molecular recognition features (MoRFs)

Several proteomic studies in the last decade revealed that many proteins are either completely disordered or possess long structurally flexible regions. Many such regions were shown to be of functional importance, often allowing a protein to interact with a large number of diverse partners. Parallel to these findings, during the last five years structural bioinformatics has produced an explosion of results regarding protein-protein interactions and their importance for cell signaling. We studied the occurrence of relatively short (10-70 residues), loosely structured protein regions within longer, largely disordered sequences that were characterized as bound to larger proteins. We call these regions molecular recognition features (MoRFs, also known as molecular recognition elements, MoREs). Interestingly, upon binding to their partner(s), MoRFs undergo disorder-to-order transitions. Thus, in our interpretation, MoRFs represent a class of disordered region that exhibits molecular recognition and binding functions. This work extends previous research showing the importance of flexibility and disorder for molecular recognition. We describe the development of a database of MoRFs derived from the RCSB Protein Data Bank and present preliminary results of bioinformatics analyses of these sequences. Based on the structure adopted upon binding, at least three basic types of MoRFs are found: α-MoRFs, β-MoRFs, and ι-MoRFs, which form α-helices, β-strands, and irregular secondary structure when bound, respectively. Our data suggest that functionally significant residual structure can exist in MoRF regions prior to the actual binding event. The contribution of intrinsic protein disorder to the nature and function of MoRFs has also been addressed. The results of this study will advance the understanding of protein-protein interactions and help towards the future development of useful protein-protein binding site predictors.     

More than just tails: intrinsic disorder in histone proteins

Many biologically active proteins are disordered as a whole, or contain long disordered regions. These intrinsically disordered proteins/regions are very common in nature, abundantly found in all organisms, where they carry out important biological functions. The functions of these proteins complement the functional repertoire of "normal" ordered proteins, and many protein functional classes are heavily dependent on intrinsic disorder. Among these disorder-centric functions are interactions with nucleic acids and protein complex assembly. In this study, we present the results of comprehensive bioinformatics analyses of the abundance and roles of intrinsic disorder in 2007 histones from 746 species. We show that all the members of the histone family are intrinsically disordered proteins. Furthermore, intrinsic disorder is not only abundant in histones, but is absolutely necessary for various histone functions, starting from heterodimerization to formation of higher order oligomers, to interactions with DNA and other proteins, and to posttranslational modifications.     

CENP-C, an autoantigen in scleroderma, is a component of the human inner kinetochore plate.

We have isolated and characterized a set of overlapping cDNA clones that encode the human centromere autoantigen centromere protein C (CENP-C). The identity of these clones has been established using several criteria. First, they were shown to encode a polypeptide that migrates at the expected position for CENP-C on SDS-polyacrylamide gel electrophoresis. Second, we have demonstrated that this polypeptide shares at least two epitopes with human CENP-C. Polyclonal antibodies were raised to fusion proteins encoded by nonoverlapping regions of the cDNA clones. These antibodies were shown to recognize a protein at a position appropriate for CENP-C on immunoblots of human chromosomal proteins. In addition, we used indirect immunofluorescence to demonstrate that these antibodies recognize centromeres of HeLa chromosomes in the expected pattern for CENP-C. Localization of CENP-C by immunoelectron microscopy reveals that this protein is a component of the inner kinetochore plate.     

Calreticulin as a new histone binding protein in mitotic chromosomes

Calreticulin (CRT) is a multifunctional Ca(2+)-binding protein that mainly functions in the endoplasmic reticulum as a molecular chaperone for newly synthesized proteins. Recently we reported the protein composition of human metaphase chromosomes (Uchiyama et al., 2004), which included CRT. Here we describe new characteristics of CRT in vitro as well as its localization on the surface of metaphase chromosomes in vivo. CRT was detected in the chromosomal fraction by Western blotting and its binding partners were identified as core and linker histones by ligand overlay assay. Surface plasmon resonance sensor analyses revealed that CRT is bound to chromatin fibers. Moreover, we found that CRT has both supercoiling activity, which assists core histone assembly into chromatin fibers, and binding ability to histone H2A/H2B dimers and histone H3/H4 tetramers. Unlike the chromosome scaffold proteins, indirect immunofluorescent staining revealed that CRT is located on the surface of metaphase chromosomes. These results suggest that CRT plays a role which involves chromatin dynamics on the surface of mitotic chromosomes.     

The evolution of unusual chromosomal systems in sciarid flies: intragenomic conflict and the sex ratio

A model is presented for the evolution of the sciarid chromosomal system. In this model, a driving X chromosome caused female-biased sex ratios. The drive was exploited by maternal autosomes that segregated with the X at spermatogenesis. Genes in mothers converted some of their XX daughters into sons by eliminating a paternal X from the embryonic soma. L chromosomes were derived from X chromosomes and favored male-biased sex ratios. An X' chromosome arose that suppressed the effects of L chromosomes. The 1:1 sex ratio is a stalemate between the X' and X'X mothers causing all-female broods and the L chromosomes in XX mothers causing all-male broods. Any element (such as an L chromosome) that is preferentially transmitted through one sex will be selected to bias the sex ratio towards this sex.     

Chromosomal divergence and evolutionary inferences in Rhodniini based on the chromosomal location of ribosomal genes

In this study, we used fluorescence in situ hybridisation to determine the chromosomal location of 45S rDNA clusters in 10 species of the tribe Rhodniini (Hemiptera: Reduviidae: Triatominae). The results showed striking inter and intraspecific variability, with the location of the rDNA clusters restricted to sex chromosomes with two patterns: either on one (X chromosome) or both sex chromosomes (X and Y chromosomes). This variation occurs within a genus that has an unchanging diploid chromosome number (2n = 22, including 20 autosomes and 2 sex chromosomes) and a similar chromosome size and genomic DNA content, reflecting a genome dynamic not revealed by these chromosome traits. The rDNA variation in closely related species and the intraspecific polymorphism in Rhodnius ecuadoriensis suggested that the chromosomal position of rDNA clusters might be a useful marker to identify recently diverged species or populations. We discuss the ancestral position of ribosomal genes in the tribe Rhodniini and the possible mechanisms involved in the variation of the rDNA clusters, including the loss of rDNA loci on the Y chromosome, transposition and ectopic pairing. The last two processes involve chromosomal exchanges between both sex chromosomes, in contrast to the widely accepted idea that the achiasmatic sex chromosomes of Heteroptera do not interchange sequences.     

Studies on Ribonucleoprotein and Argyrophilic Proteins in the Chromosomes of Prorocentrum sp.

The chromosomal ultrastructure and the chromosomal substances of ribonucleoprotein (RNP) and argyrophilic proteins of Prorocentrum sp. were studied using the techniques of conventional staining, RNP preferential staining and Ag-staining. Most part of the RNP in the chromosomes of Prorocentrum sp. was in the form of fibril distributed around the DNP throughout the chromosone. The RNP in the periphery of the chromosomes was found to form a closed membrane-like structure. Many kinds of the argyrophilic proteins were distributed regularly in the chromosome of Prorocentrum sp., but varied greatly in quantity from cell to cell. The amount of argyrophilic proteins in the chromosomes of some cells was much more than that in the nucleoplasm, but in other cells the amount of argyrophilic proteins was less than that in the nucleoplasm. The results indicated that in addition to argyrophilic proteins, RNA might play a role in the maintenance of the chromosome structure at a high level.     

A novel mammalian HORMA domain-containing protein, HORMAD1, preferentially associates with unsynapsed meiotic chromosomes

HORMA domain-containing proteins regulate interactions between homologous chromosomes (homologs) during meiosis in a wide range of eukaryotes. We have identified a mouse HORMA domain-containing protein, HORMAD1, and biochemically and cytologically shown it to be associated with the meiotic chromosome axis. HORMAD1 first accumulates on the chromosomes during the leptotene to zygotene stages of meiotic prophase I. As germ cells progress into the pachytene stage, HORMAD1 disappears from the synapsed chromosomal regions. However, once the chromosomes desynapse during the diplotene stage, HORMAD1 again accumulates on the chromosome axis of the desynapsed homologs. HORMAD1 thus preferentially localizes to unsynapsed or desynapsed chromosomal regions during the prophase I stage of meiosis. Analysis of mutant strains lacking different components of the synaptonemal complex (SC) revealed that establishment of the SC is required for the displacement of HORMAD1 from the chromosome axis. Our results therefore strongly suggest that also mammalian cells use a HORMA domain-containing protein as part of a surveillance system that monitors synapsis or other interactions between homologs.     

Null alleles for gliadin blocks in bread and durum wheat cultivars

Summary Wheat gliadin proteins are coded by clusters of genes (complex loci) located on the short arms of chromosomes of homoeologous groups 1 and 6 in bread (6x) and durum (4x) wheats. The proteins expressed by the various complex loci have been designated gliadin blocks. In a survey of accessions from the Germplasm Institute (C.N.R., Bari, Italy) collection, several different accessions have been found that lack particular blocks of proteins (null alleles). In some bread wheat accessions, seeds do not express gliadins that are coded by chromosomes 1D and 6A in normal cultivars. Similarly, some durum wheat accessions lack -gliadin components coded for by genes on chromosomes 1A and 1B. The missing proteins do not result from the absence of whole chromosomes, but may be the consequence of partial deletion of these genes at a complex locus or result from their silencing.     

Sex Chromosomes and Karyotype of the (Nearly) Mythical Creature,the Gila Monster,Heloderma suspectum (Squamata: Helodermatidae)

A wide variety of sex determination systems exist among squamate reptiles. They can therefore serve as an important model for studies of evolutionary transitions among particular sex determination systems. However, we still have only a limited knowledge of sex determination in certain important lineages of squamates. In this respect, one of the most understudied groups is the family Helodermatidae (Anguimorpha) encompassing the only two venomous species of lizards which are potentially lethal to human beings. We uncovered homomorphic ZZ/ZW sex chromosomes in the Gila monster (Heloderma suspectum) with a highly heterochromatic W chromosome. The sex chromosomes are morphologically similar to the ZZ/ZW sex chromosomes of monitor lizards (Varanidae). If the sex chromosomes of helodermatids and varanids are homologous, female heterogamety may be ancestral for the whole Anguimorpha group. Moreover, we found that the karyotype of the Gila monster consists of 2n = 36 chromosomes (14 larger metacentric chromosomes and 22 acrocentric microchromosomes). 2n = 36 is the widely distributed chromosomal number among squamates. In his pioneering works representing the only previous cytogenetic examination of the family Helodermatidae, Matthey reported the karyotype as 2n = 38 and suggested a different chromosomal morphology for this species. We believe that this was probably erroneously. We also discovered a strong accumulation of telomeric sequences on several pairs of microchromosomes in the Gila monster, which is a trait documented relatively rarely in vertebrates. These new data fill an important gap in our understanding of the sex determination and karyotype evolution of squamates.     

A Role for SUMO in meiotic chromosome synapsis

During meiotic prophase, homologous chromosomes engage in a complex series of interactions that ensure their proper segregation at meiosis I. A central player in these interactions is the synaptonemal complex (SC), a proteinaceous structure elaborated along the lengths of paired homologs. In mutants that fail to make SC, crossing over is decreased, and chromosomes frequently fail to recombine; consequently, many meiotic products are inviable because of aneuploidy. Here, we have investigated the role of the small ubiquitin-like protein modifier (SUMO) in SC formation during meiosis in budding yeast. We show that SUMO localizes specifically to synapsed regions of meiotic chromosomes and that this localization depends on Zip1, a major building block of the SC. A non-null allele of the UBC9 gene, which encodes the SUMO-conjugating enzyme, impairs Zip1 polymerization along chromosomes. The Ubc9 protein localizes to meiotic chromosomes, coincident with SUMO staining. In the zip1 mutant, SUMO localizes to discrete foci on chromosomes. These foci coincide with axial associations, where proteins involved in synapsis initiation are located. Our data suggest a model in which SUMO modification of chromosomal proteins promotes polymerization of Zip1 along chromosomes. The ubc9 mutant phenotype provides the first evidence for a cause-and-effect relationship between sumoylation and synapsis.