Doriana Misceo

Human-specific duplication and mosaic transcripts: the recent paralogous structure of chromosome 22.

In recent decades, comparative chromosomal banding, chromosome painting, and gene-order studies have shown strong conservation of gross chromosome structure and gene order in mammals. However, findings from the human genome sequence suggest an unprecedented degree of recent (<35 million years ago) segmental duplication. This dynamism of segmental duplications has important implications in disease and evolution. Here we present a chromosome-wide view of the structure and evolution of the most highly homologous duplications (> or = 1 kb and > or = 90%) on chromosome 22. Overall, 10.8% (3.7/33.8 Mb) of chromosome 22 is duplicated, with an average sequence identity of 95.4%. To organize the duplications into tractable units, intron-exon structure and well-defined duplication boundaries were used to define 78 duplicated modules (minimally shared evolutionary segments) with 157 copies on chromosome 22. Analysis of these modules provides evidence for the creation or modification of 11 novel transcripts. Comparative FISH analyses of human, chimpanzee, gorilla, orangutan, and macaque reveal qualitative and quantitative differences in the distribution of these duplications--consistent with their recent origin. Several duplications appear to be human specific, including a approximately 400-kb duplication (99.4%-99.8% sequence identity) that transposed from chromosome 14 to the most proximal pericentromeric region of chromosome 22. Experimental and in silico data further support a pericentromeric gradient of duplications where the most recent duplications transpose adjacent to the centromere. Taken together, these data suggest that segmental duplications have been an ongoing process of primate genome evolution, contributing to recent gene innovation and the dynamic transformation of genome architecture within and among closely related species.

Refinement of a chimpanzee pericentric inversion breakpoint to a segmental duplication cluster

BackgroundPericentric inversions are the most common euchromatic chromosomal differences among humans and the great apes. The human and chimpanzee karyotype differs by nine such events, in addition to several constitutive heterochromatic increases and one chromosomal fusion event. Reproductive isolation and subsequent speciation are thought to be the potential result of pericentric inversions, as reproductive boundaries form as a result of hybrid sterility.ResultsHere we employed a comparative fluorescence in situ hybridization approach, using probes selected from a combination of physical mapping, genomic sequence, and segmental duplication analyses to narrow the breakpoint interval of a pericentric inversion in chimpanzee involving the orthologous human 15q11-q13 region. We have refined the inversion breakpoint of this chimpanzee-specific rearrangement to a 600 kilobase (kb) interval of the human genome consisting of entirely duplicated material. Detailed analysis of the underlying sequence indicated that this region comprises multiple segmental duplications, including a previously characterized duplication of the alpha7 neuronal nicotinic acetylcholine receptor subunit gene (CHRNA7) in 15q13.3 and several Golgin-linked-to-PML, or LCR15, duplications.ConclusionsWe conclude that, on the basis of experimental data excluding the CHRNA7 duplicon as the site of inversion, and sequence analysis of regional duplications, the most likely rearrangement site is within a GLP/LCR15 duplicon. This study further exemplifies the genomic plasticity due to the presence of segmental duplications and highlights their importance for a complete understanding of genome evolution.

Primate chromosome evolution: Ancestral karyotypes, marker order and neocentromeres

In 1992 the Japanese macaque was the first species for which the homology of the entire karyotype was established by cross-species chromosome painting. Today, there are chromosome painting data on more than 50 species of primates. Although chromosome painting is a rapid and economical method for tracking translocations, it has limited utility for revealing intrachromosomal rearrangements. Fortunately, the use of BAC-FISH in the last few years has allowed remarkable progress in determining marker order along primate chromosomes and there are now marker order data on an array of primate species for a good number of chromosomes. These data reveal inversions, but also show that centromeres of many orthologous chromosomes are embedded in different genomic contexts. Even if the mechanisms of neocentromere formation and progression are just beginning to be understood, it is clear that these phenomena had a significant impact on shaping the primate genome and are fundamental to our understanding of genome evolution. In this report we complete and integrate the dataset of BAC-FISH marker order for human syntenies 1, 2, 4, 5, 8, 12, 17, 18, 19, 21, 22 and the X. These results allowed us to develop hypotheses about the content, marker order and centromere position in ancestral karyotypes at five major branching points on the primate evolutionary tree: ancestral primate, ancestral anthropoid, ancestral platyrrhine, ancestral catarrhine and ancestral hominoid. Current models suggest that between-species structural rearrangements are often intimately related to speciation. Comparative primate cytogenetics has become an important tool for elucidating the phylogeny and the taxonomy of primates. It has become increasingly apparent that molecular cytogenetic data in the future can be fruitfully combined with whole-genome assemblies to advance our understanding of primate genome evolution as well as the mechanisms and processes that have led to the origin of the human genome.

A dominant STIM1 mutation causes Stormorken syndrome.

Stormorken syndrome is a rare autosomal‐dominant disease with mild bleeding tendency, thrombocytopathy, thrombocytopenia, mild anemia, asplenia, tubular aggregate myopathy, miosis, headache, and ichthyosis. A heterozygous missense mutation in STIM1 exon 7 (c.910C>T; p.Arg304Trp) (NM_003156.3) was found to segregate with the disease in six Stormorken syndrome patients in four families. Upon sensing Ca2+ depletion in the endoplasmic reticulum lumen, STIM1 undergoes a conformational change enabling it to interact with and open ORAI1, a Ca2+ release‐activated Ca2+ channel located in the plasma membrane. The STIM1 mutation found in Stormorken syndrome patients is located in the coiled‐coil 1 domain, which might play a role in keeping STIM1 inactive. In agreement with a possible gain‐of‐function mutation in STIM1, blood platelets from patients were in a preactivated state with high exposure of aminophospholipids on the outer surface of the plasma membrane. Resting Ca2+ levels were elevated in platelets from the patients compared with controls, and store‐operated Ca2+ entry was markedly attenuated, further supporting constitutive activity of STIM1 and ORAI1. Thus, our data are compatible with a near‐maximal activation of STIM1 in Stormorken syndrome patients. We conclude that the heterozygous mutation c.910C>T causes the complex phenotype that defines this syndrome.

Tracking the complex flow of chromosome rearrangements from the Hominoidea Ancestor to extant Hylobates and Nomascus Gibbons by high-resolution synteny mapping

In this study we characterized the extension, reciprocal arrangement, and orientation of syntenic chromosomal segments in the lar gibbon (Hylobates lar, HLA) by hybridization of a panel of approximately 1000 human BAC clones. Each lar gibbon rearrangement was defined by a splitting BAC clone or by two overlapping clones flanking the breakpoint. A reconstruction of the synteny arrangement of the last common ancestor of all living lesser apes was made by combining these data with previous results in Nomascus leucogenys, Hoolock hoolock, and Symphalangus syndactylus. The definition of the ancestral synteny organization facilitated tracking the cascade of chromosomal changes from the Hominoidea ancestor to the present day karyotype of Hylobates and Nomascus. Each chromosomal rearrangement could be placed within an approximate phylogenetic and temporal framework. We identified 12 lar-specific rearrangements and five previously undescribed rearrangements that occurred in the Hylobatidae ancestor. The majority of the chromosomal differences between lar gibbons and humans are due to rearrangements that occurred in the Hylobatidae ancestor (38 events), consistent with the hypothesis that the genus Hylobates is the most recently evolved lesser ape genus. The rates of rearrangements in gibbons are 10 to 20 times higher than the mammalian default rate. Segmental duplication may be a driving force in gibbon chromosome evolution, because a consistent number of rearrangements involves pericentromeric regions (10 events) and centromere inactivation (seven events). Both phenomena can be reasonably supposed to have strongly contributed to the euchromatic dispersal of segmental duplications typical of pericentromeric regions. This hypothesis can be more fully tested when the sequence of this gibbon species becomes available. The detailed synteny map provided here will, in turn, substantially facilitate sequence assembly efforts.

Delineating the GRIN1 phenotypic spectrum A distinct genetic NMDA receptor encephalopathy

Objective: To determine the phenotypic spectrum caused by mutations in GRIN1 encoding the NMDA receptor subunit GluN1 and to investigate their underlying functional pathophysiology. Methods: We collected molecular and clinical data from several diagnostic and research cohorts. Functional consequences of GRIN1 mutations were investigated in Xenopus laevis oocytes. Results: We identified heterozygous de novo GRIN1 mutations in 14 individuals and reviewed the phenotypes of all 9 previously reported patients. These 23 individuals presented with a distinct phenotype of profound developmental delay, severe intellectual disability with absent speech, muscular hypotonia, hyperkinetic movement disorder, oculogyric crises, cortical blindness, generalized cerebral atrophy, and epilepsy. Mutations cluster within transmembrane segments and result in loss of channel function of varying severity with a dominant-negative effect. In addition, we describe 2 homozygous GRIN1 mutations (1 missense, 1 truncation), each segregating with severe neurodevelopmental phenotypes in consanguineous families. Conclusions: De novo GRIN1 mutations are associated with severe intellectual disability with cortical visual impairment as well as oculomotor and movement disorders being discriminating phenotypic features. Loss of NMDA receptor function appears to be the underlying disease mechanism. The identification of both heterozygous and homozygous mutations blurs the borders of dominant and recessive inheritance of GRIN1-associated disorders.

SCA27 caused by a chromosome translocation: further delineation of the phenotype

We report of a spinocerebellar ataxia (SCA)27 in a daughter and her mother whose karyotype is 46, XX t(5;13)(q31.2;q33.1). The translocation breakpoint is identical in both patients, disrupting the gene-encoding fibroblast growth factor 14 isoform b (FGF14-1b). Clinically, both show signs of SCA, although the daughter is the most affected with early onset cerebellar ataxia, microcephaly, and severe mental retardation. FGF14-1b is the predominant isoform in brain, where it interacts with the voltage gated Na channel. Fgf14−/− mice develop ataxia and paroxysmal dyskinesia and have cognitive deficits. One missense and one non-sense mutation in FGF14 have previously been linked to SCA27. Truncation of one allele in our patients suggests that haploinsuffiency of FGF14 can cause SCA27.

Spastic paraplegia type 7 is associated with multiple mitochondrial DNA deletions

Spastic paraplegia 7 is an autosomal recessive disorder caused by mutations in the gene encoding paraplegin, a protein located at the inner mitochondrial membrane and involved in the processing of other mitochondrial proteins. The mechanism whereby paraplegin mutations cause disease is unknown. We studied two female and two male adult patients from two Norwegian families with a combination of progressive external ophthalmoplegia and spastic paraplegia. Sequencing of SPG7 revealed a novel missense mutation, c.2102A>C, p.H 701P, which was homozygous in one family and compound heterozygous in trans with a known pathogenic mutation c.1454_1462del in the other. Muscle was examined from an additional, unrelated adult female patient with a similar phenotype caused by a homozygous c.1047insC mutation in SPG7. Immunohistochemical studies in skeletal muscle showed mosaic deficiency predominantly affecting respiratory complex I, but also complexes III and IV. Molecular studies in single, microdissected fibres showed multiple mitochondrial DNA deletions segregating at high levels (38–97%) in respiratory deficient fibres. Our findings demonstrate for the first time that paraplegin mutations cause accumulation of mitochondrial DNA damage and multiple respiratory chain deficiencies. While paraplegin is not known to be directly associated with the mitochondrial nucleoid, it is known to process other mitochondrial proteins and it is possible therefore that paraplegin mutations lead to mitochondrial DNA deletions by impairing proteins involved in the homeostasis of the mitochondrial genome. These studies increase our understanding of the molecular pathogenesis of SPG7 mutations and suggest that SPG7 testing should be included in the diagnostic workup of autosomal recessive, progressive external ophthalmoplegia, especially if spasticity is present.

A translocation between Xq21.33 and 22q13.33 causes an intragenic SHANK3 deletion in a woman with Phelan–McDermid syndrome and hypergonadotropic hypogonadism

Chromosome 22q13 monosomy has been described as a contiguous gene syndrome. Localized in the critical region, SHANK3 is likely to play a key role in the expression of the clinical phenotype. SHANK3 mutations have also been reported in autistic patients without a syndromic phenotype. We report on a 20‐year‐old woman with mental retardation carrying a de novo translocation between chromosome Xq21.33 and 22q13.33, associated with a duplication on Xq21.33 and deletion on 22q13.33. As a child her development was characterized by disturbed social interaction, stereotypic hand movements and ritualistic behavior and she was considered at one time to have autistic features. All these traits match the 22q13 deletion syndrome (Phelan–McDermid syndrome, OMIM 606232), likely due to the deletion overlapping the last two exons of the SHANK3 gene. Our patient harbors the smallest and most distal SHANK3 deletion described to date, yet resulting in the full spectrum of the Phelan–McDermid syndrome. In addition, she has hypergonadotropic hypogonadism with low estrogen level, high FSH level, and irregular menstruation. Intriguingly, chromosome translocations affecting the chromosome band Xq21 can result in premature ovarian failure.

A novel type of rhizomelic chondrodysplasia punctata, RCDP5, is caused by loss of the PEX5 long isoform

Import of peroxisomal matrix proteins, crucial for peroxisome biogenesis, is mediated by the cytosolic receptors PEX5 and PEX7 that recognize proteins carrying peroxisomal targeting signals 1 or 2 (PTS1 or PTS2), respectively. Mutations in PEX5 or 12 other PEX genes cause peroxisome biogenesis disorders, collectively named the Zellweger spectrum disorders (ZSDs), whereas mutations in PEX7 cause rhizomelic chondrodysplasia punctata type 1 (RCDP1). Three additional RCDP types, RCDP2-3-4, are caused, respectively, by mutations in GNPAT, AGPS and FAR1, encoding enzymes involved in plasmalogen biosynthesis. Here we report a fifth type of RCDP (RCDP5) caused by a novel mutation in PEX5. In four patients with RCDP from two independent families, we identified a homozygous frame shift mutation c.722dupA (p.Val242Glyfs(∗)33) in PEX5 (GenBank: NM_001131023.1). PEX5 encodes two isoforms, PEX5L and PEX5S, and we show that the c.722dupA mutation, located in the PEX5L-specific exon 9, results in loss of PEX5L only. Both PEX5 isoforms recognize PTS1-tagged proteins, but PEX5L is also a co-receptor for PTS2-tagged proteins. Previous patients with PEX5 mutations had ZSD, mainly due to deficient import of PTS1-tagged proteins. Similarly to mutations in PEX7, loss of PEX5L results in deficient import of PTS2-tagged proteins only, thus causing RCDP instead of ZSD. We demonstrate that PEX5L expression restores the import of PTS2-tagged proteins in patient fibroblasts. Due to the biochemical overlap between RCDP1 and RCDP5, sequencing of PEX7 and exon 9 in PEX5 should be performed in patients with a selective defect in the import of PTS2-tagged proteins.