Genome-enabled phylogenetic and functional reconstruction of an araphid pennate diatom Plagiostriata sp. CCMP470, previously assigned as a radial centric diatom, and its bacterial commensal.

Diatoms are an ecologically fundamental and highly diverse group of algae, dominating marine primary production in both open-water and coastal communities. The diatoms include both centric species, which may have radial or polar symmetry, and the pennates, which include raphid and araphid species and arose within the centric lineage. Here, we use combined microscopic and molecular information to reclassify a diatom strain CCMP470, previously annotated as a radial centric species related to Leptocylindrus danicus, as an araphid pennate species in the staurosiroid lineage, within the genus Plagiostriata. CCMP470 shares key ultrastructural features with Plagiostriata taxa, such as the presence of a sternum with parallel striae, and the presence of a highly reduced labiate process on its valve; and this evolutionary position is robustly supported by multigene phylogenetic analysis. We additionally present a draft genome of CCMP470, which is the first genome available for a staurosiroid lineage. 270 Pfams (19%) found in the CCMP470 genome are not known in other diatom genomes, which otherwise does not hold big novelties compared to genomes of non-staurosiroid diatoms. Notably, our DNA library contains the genome of a bacterium within the Rhodobacterales, an alpha-proteobacterial lineage known frequently to associate with algae. We demonstrate the presence of commensal alpha-proteobacterial sequences in other published algal genome and transcriptome datasets, which may indicate widespread and persistent co-occurrence.

Expanding the Mutation Spectrum Affecting αIIbß3 Integrin in Glanzmann Thrombasthenia: Screening of the ITGA2B and ITGB3 Genes in a Large International Cohort.

We report the largest international study on Glanzmann thrombasthenia (GT), an inherited bleeding disorder where defects of the ITGA2B and ITGB3 genes cause quantitative or qualitative defects of the αIIbß3 integrin, a key mediator of platelet aggregation. Sequencing of the coding regions and splice sites of both genes in members of 76 affected families identified 78 genetic variants (55 novel) suspected to cause GT. Four large deletions or duplications were found by quantitative real-time PCR. Families with mutations in either gene were indistinguishable in terms of bleeding severity that varied even among siblings. Families were grouped into type I and the rarer type II or variant forms with residual αIIbß3 expression. Variant forms helped identify genes encoding proteins mediating integrin activation. Splicing defects and stop codons were common for both ITGA2B and ITGB3 and essentially led to a reduced or absent αIIbß3 expression; included was a heterozygous c.1440-13_c.1440-1del in intron 14 of ITGA2B causing exon skipping in seven unrelated families. Molecular modeling revealed how many missense mutations induced subtle changes in αIIb and ß3 domain structure across both subunits, thereby interfering with integrin maturation and/or function. Our study extends knowledge of GT and the pathophysiology of an integrin.

Mutations in CERS3 cause autosomal recessive congenital ichthyosis in humans.

Autosomal recessive congenital ichthyosis (ARCI) is a rare genetic disorder of the skin characterized by abnormal desquamation over the whole body. In this study we report four patients from three consanguineous Tunisian families with skin, eye, heart, and skeletal anomalies, who harbor a homozygous contiguous gene deletion syndrome on chromosome 15q26.3. Genome-wide SNP-genotyping revealed a homozygous region in all affected individuals, including the same microdeletion that partially affects two coding genes (ADAMTS17, CERS3) and abolishes a sequence for a long non-coding RNA (FLJ42289). Whereas mutations in ADAMTS17 have recently been identified in autosomal recessive Weill-Marchesani-like syndrome in humans and dogs presenting with ophthalmologic, cardiac, and skeletal abnormalities, no disease associations have been described for CERS3 (ceramide synthase 3) and FLJ42289 so far. However, analysis of additional patients with non-syndromic ARCI revealed a splice site mutation in CERS3 indicating that a defect in ceramide synthesis is causative for the present skin phenotype of our patients. Functional analysis of patient skin and in vitro differentiated keratinocytes demonstrated that mutations in CERS3 lead to a disturbed sphingolipid profile with reduced levels of epidermis-specific very long-chain ceramides that interferes with epidermal differentiation. Taken together, these data present a novel pathway involved in ARCI development and, moreover, provide the first evidence that CERS3 plays an essential role in human sphingolipid metabolism for the maintenance of epidermal lipid homeostasis.

A novel amino acid substitution of integrin αIIb in Glanzmann thrombasthenia confirms that the N-terminal region of the receptor plays a role in maintaining ß-propeller structure.

Mutation screening in Glanzmann thrombasthenia (GT) is now advanced. Despite the large number of genetic defects reported in the ITGA2B gene, few affect the structure of the N-terminal domain of the αIIb subunit. We now report a Catalan family where type I GT is given by compound heterozygosity within ITGA2B with a Gly13Val substitution in αIIb associated with a 13 bp deletion involving the splice site of exon 15. Molecular modelling confirmed that the Gly13Val mutation interfered with the structure of the αIIb ß-propeller and confirms that a fold-back of the N-terminus to interact with residues deep within the propeller is necessary for the normal intracellular processing of the maturing αIIbß3 integrin.

Glanzmann thrombasthenia-like syndromes associated with Macrothrombocytopenias and mutations in the genes encoding the αIIbß3 integrin.

Glanzmann thrombasthenia (GT) is the most widely studied inherited disorder of platelets; it is caused by the absence of platelet aggregation due to quantitative and/or qualitative deficiencies of the αIIbß3 integrin coded by the ITGA2B and ITGB3 genes located at 17q21-23. Although platelet count and platelet volume (and morphology) are normal in classic GT, some reports have inferred a role for αIIbß3 in megakaryocytopoiesis and some novel but rare point mutations in either of the ITGA2B and ITGB3 genes have been associated with an altered platelet production and selective deficiencies in platelet function. This was brought to light by the discovery of mutations at Arg995 in αIIb and Asp723 in ß3 that lead to platelet anisotropy (increased size variation) and thrombocytopenia. Significantly, Arg995 and Asp723 form a salt linkage binding the cytoplasmic tails of αIIbß3 together keeping the integrin in a bent resting state. Mutations weakening this link (if not abolishing it) increase the activation state of αIIbß3 and interfere with megakaryocytopoiesis. Other mutations affecting platelet production involve extracellular but membrane proximal domains of ß3. Our purpose is to review the mutations in the ITGA2B and ITGB3 genes that lead to anisotropy and to discuss mechanisms by which this can be brought about.

Are bone defects in rare patients with Glanzmann's thrombasthenia associated with ITGB3 or ITGA2B mutations?

The question as to whether Glanzmann thrombasthenia patients with ITGB3 defects and deficiencies of both αIIbß3 and αvß3 show phenotypic differences to those with abnormalities exclusive to αIIbß3 is unresolved. Studies on ß3-deficient mice have shown an increased bone mass. Here we review the literature on bone defects in thrombasthenia patients and report the molecular analysis of a patient associating a lifelong thrombasthenia-like syndrome with skeletal defects. We show that the patient is compound heterozygote for Arg327His and Gly391Arg mutations in αIIb, with one mutation inherited from each parent. Modelling strongly suggested that both mutations act by destabilizing the αIIb beta propeller. So it appears likely that this patient has a combination of co-expressed genetic defects.

DNA replication origin interference increases the spacing between initiation events in human cells.

Mammalian DNA replication origins localize to sites that range from base pairs to tens of kilobases. A regular distribution of initiations in individual cell cycles suggests that only a limited number of these numerous potential start sites are converted into activated origins. Origin interference can silence redundant origins; however, it is currently unknown whether interference participates in spacing functional human initiation events. By using a novel hybridization strategy, genomic Morse code, on single combed DNA molecules from primary keratinocytes, we report the initiation sites present on 1.5 Mb of human chromosome 14q11.2. We confirm that initiation zones are widespread in human cells, map to intergenic regions, and contain sequence motifs found at other mammalian initiation zones. Origins used per cell cycle are less abundant than the potential sites of initiation, and their limited use increases the spacing between initiation events. Between-zone interference decreases in proportion to the distance from the active origin, whereas within-zone interference is 100% efficient. These results identify a hierarchical organization of origin activity in human cells. Functional origins govern the probability that nearby origins will fire in the context of multiple potential start sites of DNA replication, and this is mediated by origin interference.

Mutations in the transporter ABCA12 are associated with lamellar ichthyosis type 2.

Lamellar ichthyosis type 2 (LI2) is a rare autosomal recessive skin disorder for which a gene has been localized on chromosome 2q33-35. We report the identification of five missense mutations in the ABCA12 gene in nine families from Africa affected by LI2. The mutations were homozygous in eight consanguineous families and heterozygous in one non-consanguineous family. Four of these mutations are localized in the first ATP-binding domain (nucleotide-binding fold), which is highly conserved in all ABC proteins. The ABCA12 protein belongs to a superfamily of membrane proteins that translocate a variety of substrates across extra- and intracellular membranes. ABCA transporters have been implicated in several autosomal recessive disorders, notably of lipid metabolism. By analogy with ABCA3, a lamellar body membrane protein in lung alveolar type II cells, ABCA12 could function in cellular lipid trafficking in keratinocytes.

An integrated analysis of the genome of the hyperthermophilic archaeon Pyrococcus abyssi.

The hyperthermophilic euryarchaeon Pyrococcus abyssi and the related species Pyrococcus furiosus and Pyrococcus horikoshii, whose genomes have been completely sequenced, are presently used as model organisms in different laboratories to study archaeal DNA replication and gene expression and to develop genetic tools for hyperthermophiles. We have performed an extensive re-annotation of the genome of P. abyssi to obtain an integrated view of its phylogeny, molecular biology and physiology. Many new functions are predicted for both informational and operational proteins. Moreover, several candidate genes have been identified that might encode missing links in key metabolic pathways, some of which have unique biochemical features. The great majority of Pyrococcus proteins are typical archaeal proteins and their phylogenetic pattern agrees with its position near the root of the archaeal tree. However, proteins probably from bacterial origin, including some from mesophilic bacteria, are also present in the P. abyssi genome.

The DNA sequence and analysis of human chromosome 14.

Chromosome 14 is one of five acrocentric chromosomes in the human genome. These chromosomes are characterized by a heterochromatic short arm that contains essentially ribosomal RNA genes, and a euchromatic long arm in which most, if not all, of the protein-coding genes are located. The finished sequence of human chromosome 14 comprises 87,410,661 base pairs, representing 100% of its euchromatic portion, in a single continuous segment covering the entire long arm with no gaps. Two loci of crucial importance for the immune system, as well as more than 60 disease genes, have been localized so far on chromosome 14. We identified 1,050 genes and gene fragments, and 393 pseudogenes. On the basis of comparisons with other vertebrate genomes, we estimate that more than 96% of the chromosome 14 genes have been annotated. From an analysis of the CpG island occurrences, we estimate that 70% of these annotated genes are complete at their 5' end.