Heterozygous variants in CTR9, which encodes a major component of the PAF1 complex, are associated with a neurodevelopmental disorder.

PURPOSE: CTR9 is a subunit of the PAF1 complex (PAF1C) that plays a crucial role in transcription regulation by binding CTR9 to RNA polymerase II. It is involved in transcription-coupled histone modification through promoting H3K4 and H3K36 methylation. We describe the clinical and molecular studies in 13 probands, harboring likely pathogenic CTR9 missense variants, collected through GeneMatcher. METHODS: Exome sequencing was performed in all individuals. CTR9 variants were assessed through 3-dimensional modeling of the activated human transcription complex Pol II-DSIF-PAF-SPT6 and the PAF1/CTR9 complex. H3K4/H3K36 methylation analysis, mitophagy assessment based on tetramethylrhodamine ethyl ester perchlorate immunofluorescence, and RNA-sequencing in skin fibroblasts from 4 patients was performed. RESULTS: Common clinical findings were variable degrees of intellectual disability, hypotonia, joint hyperlaxity, speech delay, coordination problems, tremor, and autism spectrum disorder. Mild dysmorphism and cardiac anomalies were less frequent. For 11 CTR9 variants, de novo occurrence was shown. Three-dimensional modeling predicted a likely disruptive effect of the variants on local CTR9 structure and protein interaction. Additional studies in fibroblasts did not unveil the downstream functional consequences of the identified variants. CONCLUSION: We describe a neurodevelopmental disorder caused by (mainly) de novo variants in CTR9, likely affecting PAF1C function.

Human papillomavirus (HPV) contamination of gynaecological equipment.

OBJECTIVE: The gynaecological environment can become contaminated by human papillomavirus (HPV) from healthcare workers' hands and gloves. This study aimed to assess the presence of HPV on frequently used equipment in gynaecological practice. METHODS: In this cross-sectional study, 179 samples were taken from fomites (glove box, lamp of a gynaecological chair, gel tubes for ultrasound, colposcope and speculum) in two university hospitals and in four gynaecological private practices. Samples were collected with phosphate-buffered saline-humidified polyester swabs according to a standardised pattern, and conducted twice per day for 2 days. The samples were analysed by a semiquantitative real-time PCR. Statistical analysis was performed using Pearson's χ(2) test and multivariate regression analysis. RESULTS: Thirty-two (18%) HPV-positive samples were found. When centres were compared, there was a higher risk of HPV contamination in gynaecological private practices compared with hospitals (OR 2.69, 95% CI 1.06 to 6.86). Overall, there was no difference in the risk of contamination with respect to the time of day (OR 1.79, 95% CI 0.68 to 4.69). When objects were compared, the colposcope had the highest risk of contamination (OR 3.02, 95% CI 0.86 to 10.57). CONCLUSIONS: Gynaecological equipment and surfaces are contaminated by HPV despite routine cleaning. While there is no evidence that contaminated surfaces carry infectious viruses, our results demonstrate the need for strategies to prevent HPV contamination. These strategies, based on health providers' education, should lead to well-established cleaning protocols, adapted to gynaecological rooms, aimed at eliminating HPV material.

Potocki-Shaffer deletion encompassing ALX4 in a patient with frontonasal dysplasia phenotype.

Frontonasal dysplasia (FND) is a genetically heterogeneous malformation spectrum with marked hypertelorism, broad nasal tip and bifid nose. Only a small number of genes have been associated with FND phenotypes until now, the first gene being EFNB1, related to craniofrontonasal syndrome (CFNS) with craniosynostosis in addition, and more recently the aristaless-like homeobox genes ALX3, ALX4, and ALX1, which have been related with distinct phenotypes named FND1, FND2, and FND3 respectively. We here report on a female patient presenting with severe FND features along with partial alopecia, hypogonadism and intellectual disability. While molecular investigations did not reveal mutations in any of the known genes, ALX4, ALX3, ALX1 and EFNB1, comparative genomic hybridization (array CGH) techniques showed a large heterozygous de novo deletion at 11p11.12p12, encompassing the ALX4 gene. Deletions in this region have been described in patients with Potocki-Shaffer syndrome (PSS), characterized by biparietal foramina, multiple exostoses, and intellectual disability. Although the patient reported herein manifests some overlapping features of FND and PPS, it is likely that the observed phenotype maybe due to a second unidentified mutation in the ALX4 gene. The phenotype will be discussed in view of the deleted region encompassing the ALX4 gene.

Mitochondrial tRNA(Leu(UUR)) mutation m.3302A > G presenting as childhood-onset severe myopathy: threshold determination through segregation study.

Mitochondrial tRNA(Leu(UUR)) mutation m.3302A > G is associated with respiratory chain complex I deficiency and has been described as a rare cause of mostly adult-onset slowly progressive myopathy. Five families with 11 patients have been described so far; 5 of them died young due to cardiorespiratory failure. Here, we report on a segregation study in a family with an index patient who already presented at the age of 18 months with proximal muscular hypotonia, abnormal fatigability, and lactic acidosis. This early-onset myopathy was rapidly progressive. At 8 years, the patient is wheel-chair bound, requires nocturnal assisted ventilation, and suffers from recurrent respiratory infections. Severe complex I deficiency and nearly homoplasmy for m.3302A > G were found in muscle. We collected blood, hair, buccal swabs and muscle biopsies from asymptomatic adults in this pedigree and determined heteroplasmy levels in these tissues as well as OXPHOS activities in muscle. All participating asymptomatic adults had normal OXPHOS activities. In contrast to earlier reports, we found surprisingly little variation of heteroplasmy levels in different tissues of the same individual. Up to 45% mutation load in muscle and up to 38% mutation load in other tissues were found in non-affected adults. The phenotypic spectrum of tRNA(Leu(UUR)) m.3302A > G mutation seems to be wider than previously described. A threshold of more than 45% heteroplasmy in muscle seems to be necessary to alter complex I activity leading to clinical manifestation. The presented data may be helpful for prognostic considerations and counseling in affected families.

Randomized Comparison of Two Vaginal Self-Sampling Methods for Human Papillomavirus Detection: Dry Swab versus FTA Cartridge.

BACKGROUND: Human papillomavirus (HPV) self-sampling (self-HPV) is valuable in cervical cancer screening. HPV testing is usually performed on physician-collected cervical smears stored in liquid-based medium. Dry filters and swabs are an alternative. We evaluated the adequacy of self-HPV using two dry storage and transport devices, the FTA cartridge and swab. METHODS: A total of 130 women performed two consecutive self-HPV samples. Randomization determined which of the two tests was performed first: self-HPV using dry swabs (s-DRY) or vaginal specimen collection using a cytobrush applied to an FTA cartridge (s-FTA). After self-HPV, a physician collected a cervical sample using liquid-based medium (Dr-WET). HPV types were identified by real-time PCR. Agreement between collection methods was measured using the kappa statistic. RESULTS: HPV prevalence for high-risk types was 62.3% (95%CI: 53.7-70.2) detected by s-DRY, 56.2% (95%CI: 47.6-64.4) by Dr-WET, and 54.6% (95%CI: 46.1-62.9) by s-FTA. There was overall agreement of 70.8% between s-FTA and s-DRY samples (kappa = 0.34), and of 82.3% between self-HPV and Dr-WET samples (kappa = 0.56). Detection sensitivities for low-grade squamous intraepithelial lesion or worse (LSIL+) were: 64.0% (95%CI: 44.5-79.8) for s-FTA, 84.6% (95%CI: 66.5-93.9) for s-DRY, and 76.9% (95%CI: 58.0-89.0) for Dr-WET. The preferred self-collection method among patients was s-DRY (40.8% vs. 15.4%). Regarding costs, FTA card was five times more expensive than the swab (~5 US dollars (USD)/per card vs. ~1 USD/per swab). CONCLUSION: Self-HPV using dry swabs is sensitive for detecting LSIL+ and less expensive than s-FTA. TRIAL REGISTRATION: International Standard Randomized Controlled Trial Number (ISRCTN): 43310942.

Transcriptional control of Notch signaling by a HOX and a PBX/EXD protein during vulval development in C. elegans.

The Notch signaling pathway controls growth, differentiation and patterning in divergent animal phyla; in humans, defective Notch signaling has been implicated in cancer, stroke and neurodegenerative disorders. Despite its developmental and medical significance, little is known about the factors that render cells to become competent for Notch signaling. Here we show that during vulval development in the nematode Caenorhabditis elegans the HOX protein LIN-39 and its EXD/PBX-like cofactor CEH-20 are required for LIN-12/Notch-mediated lateral signaling that specifies the 2 degrees vulval cell fate. Inactivation of either lin-39 or ceh-20 resulted in the misspecification of 2 degrees vulval cells and suppressed the multivulva phenotype of lin-12(n137) gain-of-function mutant animals. Furthermore, both LIN-39 and CEH-20 are required for the expression of basal levels of the genes encoding the LIN-12/Notch receptor and one of its ligands in the vulval precursor cells, LAG-2/Delta/Serrate, rendering them competent for the subsequent lin-12/Notch induction events. Our results suggest that the transcription factors LIN-39 and CEH-20, which function at the bottom of the RTK/Ras and Wnt pathways in vulval induction, serve as major integration sites in coordinating and transmitting signals to the LIN-12/Notch cascade to regulate vulval cell fates.

The Mi-2 nucleosome-remodeling protein LET-418 is targeted via LIN-1/ETS to the promoter of lin-39/Hox during vulval development in C. elegans.

The fate of the vulval cells in Caenorhabditis elegans is specified, at least in part, through a highly conserved RTK/Ras mediated signaling cascade that negatively regulates the activity of the ETS-like transcription factor LIN-1. The Hox gene lin-39 functions downstream of both, the LIN-3/RTK/Ras pathway and LIN-1 and plays a pivotal role in controlling vulva cell competence and induction. Here we show that LET-418, a C. elegans ortholog of the human NuRD component Mi-2, negatively modulates the activity of lin-39. LET-418 interacts in vivo with specific regions in the promoter of lin-39 and this interaction depends on LIN-1. Our data provide evidence for a model in which LIN-1 recruits LET-418/Mi-2 as co-repressor to the promoter of lin-39, thereby restricting its activity to the basal levels required in the vulva precursor cells (VPCs) for normal vulval development. Thus, our data suggest that the interaction between LIN-1 and LET-418/Mi-2 may link RTK/Ras signaling with chromatin remodeling and gene expression.

A heterochromatin protein 1 homologue in Caenorhabditis elegans acts in germline and vulval development.

Proteins of the highly conserved heterochromatin protein 1 (HP1) family have been found to function in the dynamic organization of nuclear architecture and in gene regulation throughout the eukaryotic kingdom. In addition to being key players in heterochromatin-mediated gene silencing, HP1 proteins may also contribute to the transcriptional repression of euchromatic genes via the recruitment to specific promoters. To investigate the role played by these different activities in specific developmental pathways, we identified HP1 homologues in the genome of Caenorhabditis elegans and used RNA-mediated interference to study their function. We show that one of the homologues, HPL-2, is required for the formation of a functional germline and for the development of the vulva by acting in an Rb-related pathway. We suggest that, by acting as repressors of gene expression, HP1 proteins may fulfil specific functions in both somatic and germline differentiation processes throughout development.