Transcriptional changes in trichothiodystrophy cells

Mutations in three of the genes encoding the XPB, XPD and TTDA components of transcription factor TFIIH can result in the clinical phenotype of trichothiodystrophy (TTD). Different mutations in XPB and XPD can instead cause xeroderma pigmentosum (XP). The completely different features of these disorders have been attributed to TTD being a transcription syndrome. In order to detect transcriptional differences between TTD and XP cells from the XP-D complementation group, we have compared gene expression profiles in cultured fibroblasts from normal, XP and TTD donors. Although we detected transcriptional differences between individual cell strains, using an algorithm of moderate stringency, we did not identify any genes whose expression was reproducibly different in proliferating fibroblasts from each type of donor. Following UV-irradiation, many genes were up- and down-regulated in all three cell types. The microarray analysis indicated some apparent differences between the different donor types, but on more detailed inspection, these turned out to be false positives. We conclude that there are minimal differences in gene expression in proliferating fibroblasts from TTD, XP-D and normal donors

Gene expression signatures associated with chronic endometritis revealed by RNA sequencing

IntroductionChronic endometritis (CE) is a persistent inflammatory condition of the endometrium characterized by the infiltration of plasma cells in the endometrial stroma. CD138 immunohistochemistry is considered to improve the CE diagnosis rate.MethodsUsing the number of CD138-positive cells equal or greater than five as a diagnostic criterion for CE, we identified 24 CE and 33 non-CE cases among women with infertility. We conducted RNA-sequencing analysis for these 57 cases in total as an attempt to elucidate the molecular pathogenesis of CE and to search for new biomarkers for CE.Results and DiscussionBy comparing CE and non-CE groups, we identified 20 genes upregulated in the endometria of CE patients, including 12 immunoglobulin-related genes and eight non-immunoglobulin genes as differentially expressed genes. The eight genes were MUC5AC, LTF, CAPN9, MESP1, ACSM1, TVP23A, ALOX15, and MZB1. By analyzing samples in the proliferative and secretory phases of the menstrual cycle separately, we also identified four additional non-immunoglobulin genes upregulated in CE endometria: CCDC13 by comparing the samples in the proliferative phase, and OVGP1, MTUS2, and CLIC6 by comparing the samples in the secretory phase. Although the genes upregulated in CE may serve as novel diagnostic markers of CE, many of them were upregulated only in a limited number of CE cases showing an extremely high number of CD138-positive cells near or over one hundred. Exceptionally, TVP23A was upregulated in the majority of CE cases regardless of the number of CD138-positive cells. The upregulation of TVP23A in the endometria of CE cases may reflect the pathophysiology of a cell-type or cell-types intrinsic to the endometrium rather than the accumulation of plasma cells. Our data, consisting of clinical and transcriptomic information for CE and non-CE cases, helped us identify gene expression signatures associated with CE

Whole exome sequencing of circulating tumor cells provides a window into metastatic prostate cancer

Comprehensive analyses of cancer genomes promise to inform prognoses and precise cancer treatments. A major barrier, however, is inaccessibility of metastatic tissue. A potential solution is to characterize circulating tumor cells (CTCs), but this requires overcoming the challenges of isolating rare cells and sequencing low-input material. Here we report an integrated process to isolate, qualify and sequence whole exomes of CTCs with high fidelity, using a census-based sequencing strategy. Power calculations suggest that mapping of >99.995% of the standard exome is possible in CTCs. We validated our process in two prostate cancer patients including one for whom we sequenced CTCs, a lymph node metastasis and nine cores of the primary tumor. Fifty-one of 73 CTC mutations (70%) were observed in matched tissue. Moreover, we identified 10 early-trunk and 56 metastatic-trunk mutations in the non-CTC tumor samples and found 90% and 73% of these, respectively, in CTC exomes. This study establishes a foundation for CTC genomics in the clinic

Protein instability associated with AARS1 and MARS1 mutations causes trichothiodystrophy

Trichothiodystrophy (TTD) is a rare hereditary neurodevelopmental disorder defined by sulfur-deficient brittle hair and nails and scaly skin, but with otherwise remarkably variable clinical features. The photosensitive TTD (PS-TTD) forms exhibits in addition to progressive neuropathy and other features of segmental accelerated aging and is associated with impaired genome maintenance and transcription. New factors involved in various steps of gene expression have been identified for the different non-photosensitive forms of TTD (NPS-TTD), which do not appear to show features of premature aging. Here, we identify alanyl-tRNA synthetase 1 and methionyl-tRNA synthetase 1 variants as new gene defects that cause NPS-TTD. These variants result in the instability of the respective gene products alanyl- and methionyl-tRNA synthetase. These findings extend our previous observations that TTD mutations affect the stability of the corresponding proteins and emphasize this phenomenon as a common feature of TTD. Functional studies in skin fibroblasts from affected individuals demonstrate that these new variants also impact on the rate of tRNA charging, which is the first step in protein translation. The extension of reduced abundance of TTD factors to translation as well as transcription redefines TTD as a syndrome in which proteins involved in gene expression are unstable

Scalable whole-exome sequencing of cell-free DNA reveals high concordance with metastatic tumors

Whole-exome sequencing of cell-free DNA (cfDNA) could enable comprehensive profiling of tumors from blood but the genome-wide concordance between cfDNA and tumor biopsies is uncertain. Here we report ichorCNA, software that quantifies tumor content in cfDNA from 0.1× coverage whole-genome sequencing data without prior knowledge of tumor mutations. We apply ichorCNA to 1439 blood samples from 520 patients with metastatic prostate or breast cancers. In the earliest tested sample for each patient, 34% of patients have ≥10% tumor-derived cfDNA, sufficient for standard coverage whole-exome sequencing. Using whole-exome sequencing, we validate the concordance of clonal somatic mutations (88%), copy number alterations (80%), mutational signatures, and neoantigens between cfDNA and matched tumor biopsies from 41 patients with ≥10% cfDNA tumor content. In summary, we provide methods to identify patients eligible for comprehensive cfDNA profiling, revealing its applicability to many patients, and demonstrate high concordance of cfDNA and metastatic tumor whole-exome sequencing

Accumulation of TIP2;2 Aquaporin during Dark Adaptation Is Partially PhyA Dependent in Roots of Arabidopsis Seedlings

Light regulates the expression and function of aquaporins, which are involved in water and solute transport. In Arabidopsis thaliana, mRNA levels of one of the aquaporin genes, TIP2;2, increase during dark adaptation and decrease under far-red light illumination, but the effects of light at the protein level and on the mechanism of light regulation remain unknown. Numerous studies have described the light regulation of aquaporin genes, but none have identified the regulatory mechanisms behind this regulation via specific photoreceptor signaling. In this paper, we focus on the role of phytochrome A (phyA) signaling in the regulation of the TIP2;2 protein. We generated Arabidopsis transgenic plants expressing a TIP2;2-GFP fusion protein driven by its own promoter, and showed several differences in TIP2;2 behavior between wild type and the phyA mutant. Fluorescence of TIP2;2-GFP protein in the endodermis of roots in the wild-type seedlings increased during dark adaptation, but not in the phyA mutant. The amount of the TIP2;2-GFP protein in wild-type seedlings decreased rapidly under far-red light illumination, and a delay in reduction of TIP2;2-GFP was observed in the phyA mutant. Our results imply that phyA, cooperating with other photoreceptors, modulates the level of TIP2;2 in Arabidopsis roots