Authors: Grant J, Mahadevaiah SK, Khil P, Sangrithi MN, Royo H, Duckworth J, McCarrey JR, VandeBerg JL, Renfree MB, Taylor W, Elgar G, Camerini-Otero RD, Gilchrist MJ, Turner JM
Journal: Nature. 2012 Jul 12;487(7406):254-8
In female (XX) mammals, one of the two X chromosomes is inactivated to ensure an equal dose of X-linked genes with males (XY). X-chromosome inactivation in eutherian mammals is mediated by the non-coding RNA Xist. Xist is not found in metatherians (marsupials), and how X-chromosome inactivation is initiated in these mammals has been the subject of speculation for decades. Using the marsupial Monodelphis domestica, here we identify Rsx (RNA-on-the-silent X), an RNA that has properties consistent with a role in X-chromosome inactivation. Rsx is a large, repeat-rich RNA that is expressed only in females and is transcribed from, and coats, the inactive X chromosome. In female germ cells, in which both X chromosomes are active, Rsx is silenced, linking Rsx expression to X-chromosome inactivation and reactivation. Integration of an Rsx transgene on an autosome in mouse embryonic stem cells leads to gene silencing in cis. Our findings permit comparative studies of X-chromosome inactivation in mammals and pose questions about the mechanisms by which X-chromosome inactivation is achieved in eutherians.
Authors: Barnes AM, Cabral WA, Weis M, Makareeva E, Mertz EL, Leikin S, Eyre D, Trujillo C, Marini JC
Journal: Hum Mutat. 2012 Jun 20. doi: 10.1002/humu.22139. [Epub ahead of print]
Recessive osteogenesis imperfecta (OI) is caused by defects in genes whose products interact with type I collagen for modification and/or folding. We identified a Palestinian pedigree with moderate and lethal forms of recessive OI caused by mutations in FKBP10 or PPIB, which encode endoplasmic reticulum resident chaperone/isomerases FKBP65 and CyPB, respectively. In one pedigree branch, both parents carry a deletion in PPIB (c.563_566delACAG), causing lethal type IX OI in their two children. In another branch, a child with moderate type XI OI has a homozygous FKBP10 mutation (c.1271_1272delCCinsA). Proband FKBP10 transcripts are 4% of control and FKBP65 protein is absent from proband cells. Proband collagen electrophoresis reveals slight band broadening, compatible with ≈10% overmodification. Normal chain incorporation, helix folding, and collagen T(m) support a minimal general collagen chaperone role for FKBP65. However, there is a dramatic decrease in collagen deposited in culture despite normal collagen secretion. Mass spectrometry reveals absence of hydroxylation of the collagen telopeptide lysine involved in cross-linking, suggesting that FKBP65 is required for lysyl hydroxylase activity or access to type I collagen telopeptide lysines, perhaps through its function as a peptidylprolyl isomerase. Proband collagen to organics ratio in matrix is approximately 30% of normal in Raman spectra. Immunofluorescence shows sparse, disorganized collagen fibrils in proband matrix.
Authors: Zeissig S, Murata K, Sweet L, Publicover J, Hu Z, Kaser A, Bosse E, Iqbal J, Hussain MM, Balschun K, Röcken C, Arlt A, Günther R, Hampe J, Schreiber S, Baron JL, Moody DB, Liang TJ, Blumberg RS
Journal: Nat Med. 2012 Jun 17. doi: 10.1038/nm.2811
In most adult humans, hepatitis B is a self-limiting disease leading to life-long protective immunity, which is the consequence of a robust adaptive immune response occurring weeks after hepatitis B virus (HBV) infection. Notably, HBV-specific T cells can be detected shortly after infection, but the mechanisms underlying this early immune priming and its consequences for subsequent control of viral replication are poorly understood. Using primary human and mouse hepatocytes and mouse models of transgenic and adenoviral HBV expression, we show that HBV-expressing hepatocytes produce endoplasmic reticulum (ER)-associated endogenous antigenic lipids including lysophospholipids that are generated by HBV-induced secretory phospholipases and that lead to activation of natural killer T (NKT) cells. The absence of NKT cells or CD1d or a defect in ER-associated transfer of lipids onto CD1d results in diminished HBV-specific T and B cell responses and delayed viral control in mice. NKT cells may therefore contribute to control of HBV infection through sensing of HBV-induced modified self-lipids.
Authors: Lindhurst MJ, Parker VE, Payne F, Sapp JC, Rudge S, Harris J, Witkowski AM, Zhang Q, Groeneveld MP, Scott CE, Daly A, Huson SM, Tosi LL, Cunningham ML, Darling TN, Geer J, Gucev Z, Sutton VR, Tziotzios C, Dixon AK, Helliwell T, O'Rahilly S, Savage DB, Wakelam MJ, Barroso I, Biesecker LG, Semple RK
Journal: Nat Genet. 2012 Jun 24;44(8):928-33. doi: 10.1038/ng.2332
The phosphatidylinositol 3-kinase (PI3K)-AKT signaling pathway is critical for cellular growth and metabolism. Correspondingly, loss of function of PTEN, a negative regulator of PI3K, or activating mutations in AKT1, AKT2 or AKT3 have been found in distinct disorders featuring overgrowth or hypoglycemia. We performed exome sequencing of DNA from unaffected and affected cells from an individual with an unclassified syndrome of congenital progressive segmental overgrowth of fibrous and adipose tissue and bone and identified the cancer-associated mutation encoding p.His1047Leu in PIK3CA, the gene that encodes the p110α catalytic subunit of PI3K, only in affected cells. Sequencing of PIK3CA in ten additional individuals with overlapping syndromes identified either the p.His1047Leu alteration or a second cancer-associated alteration, p.His1047Arg, in nine cases. Affected dermal fibroblasts showed enhanced basal and epidermal growth factor (EGF)-stimulated phosphatidylinositol 3,4,5-trisphosphate (PIP(3)) generation and concomitant activation of downstream signaling relative to their unaffected counterparts. Our findings characterize a distinct overgrowth syndrome, biochemically demonstrate activation of PI3K signaling and thereby identify a rational therapeutic target.
Authors: Zhao F, Cannons JL, Dutta M, Griffiths GM, Schwartzberg PL
Journal: Immunity. 2012 Jun 29;36(6):1003-16
X-linked lymphoproliferative syndrome, characterized by fatal responses to Epstein-Barr virus infection, is caused by mutations affecting the adaptor SAP, which links SLAM family receptors to downstream signaling. Although cytotoxic defects in SAP-deficient T cells are documented, the mechanism remains unclear. We show that SAP-deficient murine CD8(+) T cells exhibited normal cytotoxicity against fibrosarcoma targets, yet had impaired adhesion to and killing of B cell and low-avidity T cell targets. SAP-deficient cytotoxic lymphocytes showed specific defects in immunological synapse organization with these targets, resulting in inefficient actin clearance. In the absence of SAP, signaling through the SLAM family members Ly108 and 2B4 resulted in increased recruitment of the SHP-1 phosphatase, associated with altered SHP-1 localization and decreased activation of Src kinases at the synapse. Hence, SAP and SLAM receptors regulate positive and negative signals required for organizing the T cell:B cell synapse and setting thresholds for cytotoxicity against distinct cellular targets.
Authors: Chen S, Li W
Journal: Nat Neurosci. 2012 May 27. doi: 10.1038/nn.3128
Retinal amacrine cells are thought to lack chromatic or color-selective light responses and have only a minor role in color processing. We found that a type of mammalian (Ictidomys tridecemlineatus) amacrine cell selectively carries a blue-On signal, which is received from a blue or short wavelength-sensitive (S) cone On bipolar cell. This glycinergic inhibitory S-cone amacrine cell is ideally placed for driving blue-Off responses in downstream ganglion cells.
Authors: Reid RJ, McBride CM, Alford SH, Price C, Baxevanis AD, Brody LC, Larson EB
Journal: Genet Med. 2012 May 17. doi: 10.1038/gim.2012.52
Purpose: The objective of this work was to examine whether offers of multiplex genetic testing increase health-care utilization among healthy patients aged 25-40 years. The identification of genetic variants associated with common disease is accelerating rapidly. "Multiplex tests" that give individuals feedback on large panels of genetic variants have proliferated. Availability of these test results may prompt consumers to use more health-care services.
Methods: A total of 1,599 continuously insured adults aged 25-40 years were surveyed and offered a multiplex genetic susceptibility test for eight common health conditions. Health-care utilization from automated records was compared in 12-month pre- and posttest periods among persons who completed a baseline survey only (68.7%), those who visited a study website but opted not to test (17.8%), and those who chose the multiplex genetic susceptibility test (13.6%).
Results: In the pretest period, persons choosing genetic testing used an average of 1.02 physician visits per quarter as compared with 0.93 and 0.82 for the baseline-only and Web-only groups, respectively (P < 0.05). There were no statistically significant differences by group in the pretest use of any common medical tests or procedures associated with four common health conditions. When changes in physician and medical test/procedure use in the posttest period were compared among the groups, no statistically significant differences were observed for any utilization category.
Conclusions: Persons offered and completing multiplex genetic susceptibility testing used more physician visits before testing, but testing was not associated with subsequent changes in use. This study supports the supposition that multiplex genetic testing offers can be provided directly to the patients in such a way that use of health services is not inappropriately increased.
Authors: Califf RM, Zarin DA, Kramer JM, Sherman RE, Aberle LH, Tasneem A
Journal: JAMA. 2012 May 2;307(17):1838-47. doi: 10.1001/jama.2012.3424
CONTEXT: Recent reports highlight gaps between guidelines-based treatment recommendations and evidence from clinical trials that supports those recommendations. Strengthened reporting requirements for studies registered with ClinicalTrials.gov enable a comprehensive evaluation of the national trials portfolio. OBJECTIVE: To examine fundamental characteristics of interventional clinical trials registered in the ClinicalTrials.gov database. METHODS: A data set comprising 96,346 clinical studies from ClinicalTrials.gov was downloaded on September 27, 2010, and entered into a relational database to analyze aggregate data. Interventional trials were identified and analyses were focused on 3 clinical specialties-cardiovascular, mental health, and oncology-that together encompass the largest number of disability-adjusted life-years lost in the United States. MAIN OUTCOME MEASURES: Characteristics of registered clinical trials as reported data elements in the trial registry; how those characteristics have changed over time; differences in characteristics as a function of clinical specialty; and factors associated with use of randomization, blinding, and data monitoring committees (DMCs). RESULTS: The number of registered interventional clinical trials increased from 28,881 (October 2004-September 2007) to 40,970 (October 2007-September 2010), and the number of missing data elements has generally declined. Most interventional trials registered between 2007 and 2010 were small, with 62% enrolling 100 or fewer participants. Many clinical trials were single-center (66%; 24,788/37,520) and funded by organizations other than industry or the National Institutes of Health (NIH) (47%; 17,592/37,520). Heterogeneity in the reported methods by clinical specialty; sponsor type; and the reported use of DMCs, randomization, and blinding was evident. For example, reported use of DMCs was less common in industry-sponsored vs NIH-sponsored trials (adjusted odds ratio [OR], 0.11; 95% CI, 0.09-0.14), earlier-phase vs phase 3 trials (adjusted OR, 0.83; 95% CI, 0.76-0.91), and mental health trials vs those in the other 2 specialties. In similar comparisons, randomization and blinding were less frequently reported in earlier-phase, oncology, and device trials. CONCLUSION: Clinical trials registered in ClinicalTrials.gov are dominated by small trials and contain significant heterogeneity in methodological approaches, including reported use of randomization, blinding, and DMCs.
Authors: Roberts SA, Sterling J, Thompson C, Harris S, Mav D, Shah R, Klimczak LJ, Kryukov GV, Malc E, Mieczkowski PA, Resnick MA, Gordenin DA
Journal: Mol Cell. 2012 May 25;46(4):424-35
Mutations are typically perceived as random, independent events. We describe here nonrandom clustered mutations in yeast and in human cancers. Genome sequencing of yeast grown under chronic alkylation damage identified mutation clusters that extend up to 200 kb. A predominance of "strand-coordinated" changes of either cytosines or guanines in the same strand, mutation patterns, and genetic controls indicated that simultaneous mutations were generated by base alkylation in abnormally long single-strand DNA (ssDNA) formed at double-strand breaks (DSBs) and replication forks. Significantly, we found mutation clusters with analogous features in sequenced human cancers. Strand-coordinated clusters of mutated cytosines or guanines often resided near chromosome rearrangement breakpoints and were highly enriched with a motif targeted by APOBEC family cytosine-deaminases, which strongly prefer ssDNA. These data indicate that hypermutation via multiple simultaneous changes in randomly formed ssDNA is a general phenomenon that may be an important mechanism producing rapid genetic variation.
Authors: Brick K, Smagulova F, Khil P, Camerini-Otero RD, Petukhova GV
Journal: Nature. 2012 May 13;485(7400):642-5. doi: 10.1038/nature11089
Genetic recombination occurs during meiosis, the key developmental programme of gametogenesis. Recombination in mammals has been recently linked to the activity of a histone H3 methyltransferase, PR domain containing 9 (PRDM9), the product of the only known speciation-associated gene in mammals. PRDM9 is thought to determine the preferred recombination sites--recombination hotspots--through sequence-specific binding of its highly polymorphic multi-Zn-finger domain. Nevertheless, Prdm9 knockout mice are proficient at initiating recombination. Here we map and analyse the genome-wide distribution of recombination initiation sites in Prdm9 knockout mice and in two mouse strains with different Prdm9 alleles and their F(1) hybrid. We show that PRDM9 determines the positions of practically all hotspots in the mouse genome, with the exception of the pseudo-autosomal region (PAR)--the only area of the genome that undergoes recombination in 100% of cells. Surprisingly, hotspots are still observed in Prdm9 knockout mice, and as in wild type, these hotspots are found at H3 lysine 4 (H3K4) trimethylation marks. However, in the absence of PRDM9, most recombination is initiated at promoters and at other sites of PRDM9-independent H3K4 trimethylation. Such sites are rarely targeted in wild-type mice, indicating an unexpected role of the PRDM9 protein in sequestering the recombination machinery away from gene-promoter regions and other functional genomic elements.