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The O-GlcNAc transferase (OGT) modifies nuclear and cytoplasmic proteins with β-N-acetyl-glucosamine (O-GlcNAc). With thousands of O-GlcNAc-modified proteins but only one OGT encoded in the mammalian genome, a prevailing question is how OGT selects its substrates. Prior work has indicated that the tetratricopeptide repeat (TPR) domain of OGT is involved in substrate selection. Furthermore, several variants of OGT causal for X-linked intellectual disability (XLID) occur in the TPR domain. Therefore, we adapted the BioID labeling method to identify interactors of a TPR-BirA* fusion protein in HeLa cells. We identified 115 interactors representing known and novel O-GlcNAc-modified proteins and OGT interactors (raw data deposited in MassIVE, Dataset ID MSV000085626). The interactors are enriched in known OGT processes (e.g., chromatin remodeling) as well as processes in which OGT has yet to be implicated (e.g., pre-mRNA processing). Importantly, the identified TPR interactors are linked to several disease states but most notably are enriched in pathologies featuring intellectual disability that may underlie the mechanism by which mutations in OGTlead to XLID. This interactome for the TPR domain of OGT serves as a jumping-off point for future research exploring the role of OGT, the TPR domain, and its protein interactors in multiple cellular processes and disease mechanisms, including intellectual disability.

Intellectual disability (ID) is a neurodevelopmental condition that affects ~1% of the world population. In total 5−10% of ID cases are due to variants in genes located on the X chromosome. Recently, variants in OGThave been shown to co-segregate with X-linked intellectual disability (XLID) in multiple families. OGTencodes O-GlcNAc transferase (OGT), an essential enzyme that catalyses O-linked glycosylation with β-N-acetylglucosamine (O-GlcNAc) on serine/threonine residues of thousands of nuclear and cytosolic proteins. In this review, we compile the work from the last few years that clearly delineates a new syndromic form of ID, which we propose to classify as a novel Congenital Disorder of Glycosylation (OGT-CDG). We discuss potential hypotheses for the underpinning molecular mechanism(s) that provide impetus for future research studies geared towards informed interventions.

•A critical knowledge gap exists regarding quantitative bear bone microarchitectural data.•Ca.N is significantly different between black bear and human metacarpals/pooled metapodials.•Osteon banding and resorption spaces are more prevalent in bear metapodials.•3D analysis of bear cortical bone provides an accurate means to infer species origin.•3D data reveal the usefulness of desktop µCT as a diverse tool for anthropologists.

ABSTRACTWe have developed and validated a nested real-time PCR (NRT-PCR) for the genotyping of Chlamydia trachomatisand used it specifically for the typing of either eight genovars from D to K or three genovars of lymphogranuloma venereum (LGV). The 11 probes used in the NRT-PCR correctly identified the DNA from D to K and LGV reference strains and did not cross-react with the DNA from 26 strains representing the bacterial pathogens and commensals of the oropharynx, genital tract, and rectum. The NRT-PCR had a 95% probability of detection at four genome copies (confidence interval, three to six copies) of C. trachomatisper reaction. One hundred cervical and urethral swab specimens containing C. trachomatisDNA from 63 women and 37 men were used to validate the method. The results from the NRT-PCR and the DNA sequencing of amplicons generated from the omp1gene showed 100% correlation for these samples. The assay also identified the LGV-II genotype in 24 of 48 rectal swab specimens containing C. trachomatisDNA that were obtained from men having sex with men. The Sexually Transmitted Bacteria Reference Laboratory, London, independently confirmed these results using group-specific LGV real-time PCR and restriction fragment length polymorphism analysis. Compared with the NRT-PCR, non-NRT-PCR was found to be less sensitive: it typed C. trachomatisDNA in only 80% of the genital samples and 90% of the rectal swab samples. This is the first successful demonstration of the use of real-time PCR for the genotype-specific typing of C. trachomatisstrains that cause sexually transmitted diseases.

ABSTRACTA multitarget real-time PCR (MRT-PCR) for detection of Chlamydia trachomatisDNA was developed and validated. There were three targets for amplification in a single reaction: the cryptic plasmid (CP), the major outer membrane protein (MOMP) gene, and an internal control. The assay had the following characteristics: (i) detection and confirmation of the presence of C. trachomatisDNA in a single reaction, (ii) detection of all genovars of C. trachomatiswithout any cross-reactivity with pathogenic bacteria or commensal organisms of the oropharynx and genital tract, (iii) a 95% probability of detection with three copies of MOMP and one copy of CP per reaction mixture, (iv) identification of the inhibition of amplification, (v) a quantitative dynamic range of 25 to 250,000 genome copies per reaction mixture, (vi) high intra- and interassay reproducibilities, and (vii) correct identification of all samples in the validation panel. There were 146 COBAS Amplicor PCR (Amplicor PCR)-positive samples and 122 Amplicor PCR-negative samples in the panel. MRT-PCR detected CP DNA alone in 6 (4%) Amplicor PCR-positive samples and both CP and MOMP DNAs in 140 (96%) of 146 Amplicor PCR-positive samples. The quantity of MOMP DNA in 95 (68%) of 140 samples was within the dynamic range of the assay. The median C. trachomatisload in these samples was 321 genome copies per reaction mixture (range, 26 to 40,137 genome copies per reaction mixture). Due to the inclusion of two different C. trachomatis-specific targets, the assay confirmed 259 (97%) of 268 results in a single reaction. This assay could be used in the qualitative format for the routine detection of C. trachomatisand in the quantitative format for study of the pathogenesis of C. trachomatis-associated diseases. The assay demonstrated the potential to eliminate the need for confirmatory testing in almost all samples, thus reducing the turnaround time and the workload.