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Orientadores: Louis Bernard Klaczko, Ana Maria L. Azeredo-Espin Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia Resumo: O grupo tripunctata e o segundo maior grupo neotropical de Drosophila em numero de espécies e foi incluído na radiação tripunctata ¿ que inclui outros grupos próximos. O segundo cromossomo de Drosophila medipunctata (espécie que pertence ao grupo tripunctata) e altamente polimorfico para inversões. Trabalhos anteriores sugeriram a presença de interação genotipo-ambiente para tamanho da asa, mas não para forma. Experimentos de laboratório foram realizados para testar os efeitos combinados de temperatura e inversão cromossômica no tamanho e forma da asa de D. mediopuncta. A morfologia da asa foi analisada por métodos de morfometria geométrica. Os resultados mostraram que tamanho e forma da asa são influenciados por temperatura, sexo e cariótipo. Também foram encontradas evidencias que sugeriram a existência entre os efeitos de cariótipo e temperatura para forma da asa, mas nao para tamanho, indicando a presença de interação genotipo-ambiente para forma da asa em D. mediopunctata. Sugerimos que outros fatores ecológicos ¿ tais como densidade de larvas ¿ ou variação sazonal de conteúdo genético das inversões poderia explicar os resultados anteriores. Alem disso, uma nova hipótese filogenética para a radiação tripunctata e sugerida com base em dados de seqüências dos genes mitocôndrias das subunidades I e II da citocromo oxidase, e a variação de tamanho e forma da asa e descrita com bases na nova hipótese filogenética. As arvores filogenéticas foram reconstruídas por métodos de parcimônia, máxima verossimilhança e inferência Bayesiana. Os resultados rejeitaram a hipótese de monofila para o grupo tripunctata, enquanto esta hipótese não foi rejeitada para a radiação tripunctata e outros grupos específicos dentro da radiação. Ambos tamanho e forma da asa apresentaram sinal filogenético e diferentes padrões de tamanho e forma puderam ser identificados para cada um dos agrupamentos mais importantes detectados pela analise filogenética Abstract: The tripunctata group is the second largest Neotropical group of Drosophila in number of species and was included in the tripunctata radiation ¿ which comprises other closely related groups. The second chromosome of Drosophila mediopunctata (a species that belongs to the tripunctata group) is highly polymorphic for inversions. Previous work suggested the presence of genotype-environment interaction for wing size but not for shape. We performed experiments in the laboratory to test for the joint effects of temperature and chromosome inversions on size and shape of the wing in D. mediopunctata. Wing morphology was analyzed by methods of geometric morphometrics. Our findings show that wing size and shape are influenced by temperature, sex, and karyotype. We also found evidence suggestive of an interaction between the effects of karyotype and temperature on wing shape (but not for size), indicating the existence of genotype-environment interaction for wing shape in D. mediopunctata. We suggest that other ecological factors ¿ such as larval crowding ¿ or seasonal variation of genetic content within inversions may explain the previous results. Moreover, we suggest a new phylogenetic hypothesis for the tripunctata radiation based on sequences of mitochondrial genes of cytochrome oxidase subunits I and II, and describe wing size and shape variation in different species based on this new phylogenetic hypothesis. Phylogenetic trees were reconstructed by parsimony, maximum likelihood and bayesian inference. Results reject the monophyly hypothesis for the tripunctata group whereas monophyly is not rejected for the tripunctata radiation and other specific groups within the radiation. Both wing size and shape displayed phylogenetic signal and different patterns of size and shape could be identified for each of the most important clusters detected by phylogenetic analysis Doutorado Genética Animal e Evolução Doutor em Genética e Biologia Molecular

Human chromosome 2 contains large blocks of segmental duplications (SDs), both within and between proximal 2p and proximal 2q, and these may contribute to the frequency of the common variant inversion inv(2)(p11.2q13). Despite their being cytogenetically homogeneous, we have identified four different breakpoint combinations by fluorescence in situ hybridization mapping of 40 cases of inv(2)(p11.2q13) of European origin. For the vast majority of inversions (35/40), the breakpoints fell within the same spanning BACs, which hybridized to both 2p11.2 and 2q13 on the normal and inverted homologues. Sequence analysis revealed that these BACs contain a significant proportion of intrachromosomal SDs with sequence homology to the reciprocal breakpoint region. In contrast, BACs spanning the rare breakpoint combinations contain fewer SDs and with sequence homology only to the same chromosome arm. Using haplotype analysis, we identified a number of related family subgroups with identical or very closely related haplotypes. However, the majority of cases were not related, demonstrating for the first time that the inv(2)(p11.2q13) is a truly recurrent rearrangement. Therefore, there are three explanations to account for the frequent observation of the inv(2)(p11.2q13): the majority have arisen independently in different ancestors, while a minority either have been transmitted from a common founder or have different breakpoints at the molecular cytogenetic level. Udgivelsesdato: 2007-Oct

Udgivelsesdato: 2006-Apr-27 After the completion of a draft human genome sequence, the International Human Genome Sequencing Consortium has proceeded to finish and annotate each of the 24 chromosomes comprising the human genome. Here we describe the sequencing and analysis of human chromosome 3, one of the largest human chromosomes. Chromosome 3 comprises just four contigs, one of which currently represents the longest unbroken stretch of finished DNA sequence known so far. The chromosome is remarkable in having the lowest rate of segmental duplication in the genome. It also includes a chemokine receptor gene cluster as well as numerous loci involved in multiple human cancers such as the gene encoding FHIT, which contains the most common constitutive fragile site in the genome, FRA3B. Using genomic sequence from chimpanzee and rhesus macaque, we were able to characterize the breakpoints defining a large pericentric inversion that occurred some time after the split of Homininae from Ponginae, and propose an evolutionary history of the inversion.

Williams-Beuren syndrome (WBS) is most often caused by hemizygous deletion of a 1.5-Mb interval encompassing at least 17 genes at 7q11.23 (refs. 1,2). As with many other haploinsufficiency diseases, the mechanism underlying the WBS deletion is thought to be unequal meiotic recombination, probably mediated by the highly homologous DNA that flanks the commonly deleted region. Here, we report the use of interphase fluorescence in situ hybridization (FISH) and pulsed-field gel electrophoresis (PFGE) to identify a genomic polymorphism in families with WBS, consisting of an inversion of the WBS region. We have observed that the inversion is hemizygous in 3 of 11 (27%) atypical affected individuals who show a subset of the WBS phenotypic spectrum but do not carry the typical WBS microdeletion. Two of these individuals also have a parent who carries the inversion. In addition, in 4 of 12 (33%) families with a proband carrying the WBS deletion, we observed the inversion exclusively in the parent transmitting the disease-related chromosome. These results suggest the presence of a newly identified genomic variant within the population that may be associated with the disease. It may result in predisposition to primarily WBS-causing microdeletions, but may also cause translocations and inversions., link_to_OA_fulltext

The synthesis of adenine and thymine cyclopentylethyl nucleosides is presented. This novel constrained monomeric building block is very difficult to incorporate into oligonucleotides. It was introduced in 13mer oligodeoxynucleotide sequences at a single position using H-phosphonate chemistry. Phosphoramidite chemistry completely failed in this particular case. The H-phosphonate building blocks were obtained starting from the corresponding phosphoramidites. Stability of duplexes with RNA and DNA is significantly reduced. ispartof: Nucleic acids research vol:25 issue:15 pages:3034-3041 ispartof: location:England status: published

Reelin is an extracellular matrix-associated protein important in the regulation of neuronal migration during cerebral cortical development. Point mutations in the RELN gene have been shown to cause an autosomal recessive human brain malformation termed lissencephaly with cerebellar hypoplasia (LCH). Recent work has raised the possibility that reelin may also play a pathogenic role in other neuropsychiatric disorders. We sought, therefore, to define more precisely the phenotype of RELN gene disruption. To do this, we performed a clinical, radiological, and molecular study of a family in whom multiple individuals carry a chromosomal inversion that disrupts the RELN locus. A 6-year-old girl homozygous for the pericentric inversion 46,XX,inv7(p11.2q22) demonstrated the same clinical features that have been previously described in association with RELN point mutations. The girl's brain magnetic resonance imaging (MRI) findings, including pachygyria and severe cerebellar hypoplasia, were identical to those seen with RELN point mutations. Fluorescence in situ hybridization confirmed that one of the breakpoints of this inversion mapped to within the RELN gene, and Western blotting revealed an absence of detectable serum reelin protein. Several relatives who were heterozygous for this inversion were neurologically normal and had no signs of psychotic illness. Our findings demonstrate the distinctive phenotype of LCH, which is easily distinguishable from other forms of lissencephaly. Although RELN appears to be critical for normal cerebral and cerebellar development, its role, if any, in the pathogenesis of psychiatric disorders remains unclear. © 2006 Wiley-Liss, Inc.

Reelin is an extracellular matrix-associated protein important in the regulation of neuronal migration during cerebral cortical development. Point mutations in the RELN gene have been shown to cause an autosomal recessive human brain malformation termed lissencephaly with cerebellar hypoplasia (LCH). Recent work has raised the possibility that reelin may also play a pathogenic role in other neuropsychiatric disorders. We sought, therefore, to define more precisely the phenotype of RELN gene disruption. To do this, we performed a clinical, radiological, and molecular study of a family in whom multiple individuals carry a chromosomal inversion that disrupts the RELN locus. A 6-year-old girl homozygous for the pericentric inversion 46,XX,inv7(p11.2q22) demonstrated the same clinical features that have been previously described in association with RELN point mutations. The girl's brain magnetic resonance imaging (MRI) findings, including pachygyria and severe cerebellar hypoplasia, were identical to those seen with RELN point mutations. Fluorescence in situ hybridization confirmed that one of the breakpoints of this inversion mapped to within the RELN gene, and Western blotting revealed an absence of detectable serum reelin protein. Several relatives who were heterozygous for this inversion were neurologically normal and had no signs of psychotic illness. Our findings demonstrate the distinctive phenotype of LCH, which is easily distinguishable from other forms of lissencephaly. Although RELN appears to be critical for normal cerebral and cerebellar development, its role, if any, in the pathogenesis of psychiatric disorders remains unclear. © 2006 Wiley-Liss, Inc.

Udgivelsesdato: 2005-Oct The X-linked form of Alport syndrome (AS) is caused by mutation in the COL4A5 gene located at Xq22.3 and encoding the alpha5-chain of type IV-collagen. More than 400 different mutations have so far been detected in the COL4A5 gene. Not all mutations, however, will be detected using an exon-by-exon mutation detection strategy such as SSCP analysis or direct sequencing. We have previously reported the results of SSCP analysis of 81 patients suspected of X-linked AS. Genomic DNA from these 81 patients was also analyzed for larger genomic rearrangements, using Southern blotting analysis. Abnormal band patterns were found in three patients, two of which were caused by single base substitutions in the coding region, also detected by the SSCP analysis. Here we report the results of the analysis of a larger structural COL4A5 rearrangement that escaped the SSCP analysis. The rearrangement was found to be an inversion of a 21 Mb fragment of the COL4A5 gene comprising exon 9 through 51 with proximal breakpoint within intron 8 at Xq22.3 and a distal breakpoint 56 kb upstream to the initiation codon in the RAB33A gene at Xq25. The inversion of exon 9 through 51 is expected to result in a truncated or absent alpha5(IV)-chain and has not previously been associated with AS. These findings emphasize the need for a supplement to mutation detection strategies such as SSCP analysis and direct sequencing, in order to detect more complicated structural COL4A5 rearrangements. Larger structural rearrangements constitute 2.3% (1/43) of the mutations in the present material.

A dysmorphic newborn with 45,x,der(1)inv,(1)(p13;qter)t(y;1)(pter -> q11;p13),-Y de novo karyotype: Y/autosome translocations are very rare chromosomal rearrangements. In most cases, the long arm of the Y chromosome is translocated onto an autosome and most patients are referred because of male infertility. Y/1 translocations are very rare, and have been reported in seven patients sofar. Pericentric inversions may be seen In all chromosomes and are not associated with phenotypic abnormalities. Here we report a 6-day old male baby with prenatal growth retardation, frontal bossing, hypertelorism. micrognathia, cleft soft palate, absent uvula, hypospadias, simian line in both hands and hammer toes. Cytogenetic analysis was performed with GTG-banding, C-banding and FISH analysis containing X centromeric probe, Yq12-qter locus specific probe and whole chromosome Y probe. An unbalanced Y/1 translocation was diagnosed: 45,X,der(1)inv(1)(p13:qter)t(y;1)(pter -> q11:p13),-Y.

Background. Cytogenetic studies in patients with reproductive failure Aim. To investigate the contribution of chromosomal abnormalities in sub fertility and in couples with repeated abortions. Methods. Hundred and 13 couples who had at least two or more spontaneous abortions and 65 women and 63 men with infertility were analyzed cytogenetically. Results. Major chromosomal rearrangements were found in 8% and minor variants in 6% in the study population. Major chromosomal aberrations were judged to explain 4.9% of recurrent abortions and 13% of infertility. Chromosomal abnormalities in infertile men occurred in 5% and in infertile women in 21.5%. The chromosomal abnormalities were structural (57%), numerical (18%) or mosaics (25%). Conclusions. Chromosomal aberrations in recurrent abortions are mostly structural ones and those in female infertility mosaicism of sex chromosomes. Turner's syndrome, Turner variants and XY females are detected as a cause of female infertility. The structural and numerical aberrations of either sex or autosomal chromosomes were found in infertile men.

The cactophilic Drosophila buzzatii provides an excellent model for the study of reaction norms across discrete environments because it breeds on rotting tissues (rots) of very different cactus species. Here we test the possible effects of second chromosome inversions on body size and shape (wing loading) across suitable natural breeding substrates. Using homokaryotypic stocks derived from several lines homozygous for four naturally occurring chromosomal inversions, we show that arrangements significantty affect size-related traits and wing loading. In addition, karyotypes show differing effects, across natural breeding resources, for wing loading. The 2st and 2jz3 arrangements decrease and the 2j arrangement increases wing loading. For thorax length and wing loading, karyotypic correlations across host plants are slightly lower in females than in males. These results support the hypothesis that these traits have a genetic basis associated with the inversion polymorphism. Fil:Fernández Iriarte, P.J. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Norry, F.M. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Hasson, E.R. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.

The inversion polymorphism of the cactophilic fly Drosophila buzzatii was studied in two natural populations. We assessed the temporal changes and microspatial population structure. We observed a significant increase in the frequency of arrangement 2J at the expense of 2ST in both populations. These gene arrangements appear to affect the life-history of flies differently. Environmental heterogeneity explains the karyotype coexistence in nature. The analysis of population structure showed that differentiation of inversion frequencies among individual breeding sites, the rotting clacodes of Opuntia vulgaris, was highly significant. The karyotypic frequencies did not depart significantly from Hardy-Weinberg expectations, neither in individual rots nor in the total population. These results suggest that the observed population structure can be easily accounted by random genetic drift. Fil:Fernández Iriarte, P.J. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Levy, E. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Rodríguez, C. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Fanara, J.J. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Hasson, E. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.

Previous works in a colonized and an original population of Drosophila buzzatii have shown a consistent relationship between the inversion polymorphism and thorax length, a measure of body size. However, the populations studied in those reports share a close genealogical relationship as suggested by several lines of evidence. In the present paper, we revisit this issue by analysing the correlation between second chromosome arrangements and thorax length in two Argentinian natural populations (Termas de Rio Hondo and Arroyo Escobar) from different biogeographic areas with different host plants. Our findings are: (1) inversion frequencies were significantly different between populations; (2) the mean thorax length of flies collected in both populations was not significantly different; and (3) we obtain confirming evidence that flies carrying 2st, the ancestral gene order, have on average a smaller body size than those carrying the derived arrangements (2j and 2jz3). These results suggest that the biometrical effect of inversions on body size previously described are due to genetic differences between arrangements and not to the close historical relationship between the populations studied in previous reports. Fil:Fanara, J.J. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Hasson, E. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Rodríguez, C. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.

The most-dominant surface-exposed protein in many bacterial species is the S-protein. This protein crystallises into a regular monolayer on the outside surface of the bacteria: the S-layer. Lactobacillus acidophilus harbours two S-protein-encoding genes, slpA and slpB, only one of which (slpA) is expressed. In this study, we show by polymerase chain reaction (PCR) analysis that slpA and slpB are located on a 6 kb chromosomal segment, in opposite or orientations. In a small fraction of the bacterial population, this segment is inverted. The inversion leads to interchanging of the expressed and silent S-protein-encoding genes, and places the formerly silent gene behind the S-promoter which is located outside the inverted segment. A 26 bp sequence showing a high degree of similarity with the consensus sequence recognized by the Din family of invertases is present in the region where recombination occurs. Expression of the slpA gene seems to be favoured under laboratory growth conditions because 99.7% of the chromosomes of an L. acidophilus ATCC 4356 broth culture had the slpA gene present at the slp expression site.