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The ubiquitin‐proteasome proteolytic pathway is pivotal in most biological processes. Despite a great level of information available for the eukaryotic 26S proteasome—the protease responsible for the degradation of ubiquitylated proteins—several structural and functional questions remain unanswered. To gain more insight into the assembly and function of the metazoan 26S proteasome, a two‐hybrid‐based protein interaction map was generated using 30 Caenorhabditis elegans proteasome subunits. The results recapitulate interactions reported for other organisms and reveal new potential interactions both within the 19S regulatory complex and between the 19S and 20S subcomplexes. Moreover, novel potential proteasome interactors were identified, including an E3 ubiquitin ligase, transcription factors, chaperone proteins and other proteins not yet functionally annotated. By providing a wealth of novel biological hypotheses, this interaction map constitutes a framework for further analysis of the ubiquitin‐proteasome pathway in a multicellular organism amenable to both classical genetics and functional genomics.

The genome sequences of Caenorhabditis elegans , Drosophila melanogaster and Arabidopsis thaliana have been predicted to contain 19,000, 13,600 and 25,500 genes, respectively 1 , 2 , 3 . Before this information can be fully used for evolutionary and functional studies, several issues need to be addressed. First, the gene number estimates obtained in silico and not yet supported by any experimental data need to be verified. For example, it seems biologically paradoxical that C. elegans would have 50% more genes than Drosophilia . Second, intron/exon predictions need to be tested experimentally. Third, complete sets of open reading frames (ORFs), or “ORFeomes,” 4 need to be cloned into various expression vectors. To address these issues simultaneously, we have designed and applied to C. elegans the following strategy. Predicted ORFs are amplified by PCR from a highly representative cDNA library 4 using ORF-specific primers, cloned by Gateway recombination cloning 4 , 5 , 6 and then sequenced to generate ORF sequence tags (OSTs) as a way to verify identity and splicing. In a sample (n=1,222) of the nearly 10,000 genes predicted ab initio (that is, for which no expressed sequence tag (EST) is available so far), at least 70% were verified by OSTs. We also observed that 27% of these experimentally confirmed genes have a structure different from that predicted by GeneFinder. We now have experimental evidence that supports the existence of at least 17,300 genes in C. elegans . Hence we suggest that gene counts based primarily on ESTs may underestimate the number of genes in human and in other organisms.

Itamar Rabinovich (Op-Ed, Sept. 1) asserts that opportunities for peace in the Middle East were squandered by Syria's insistence that Israel formally commit to withdrawal from the Golan to the pre-June 4, 1967, borders as a precondition to a resumption of negotiations.

The first wave of COVID-19 was accompanied by global uncertainty. Delayed presentation of patients to hospitals ensued, with surgical pathologies no exception. This study aimed to assess whether delayed presentations resulted in more complex appendicectomies during the first wave of COVID-19. Operation notes for all presentations of appendicitis ( =216) within a single health board (three hospitals) during two three-month periods (control period (pre-COVID) vs COVID pandemic) were analysed, and the severity of appendicitis was recorded as per the American Association for the Surgery of Trauma (AAST) grading system. Presentations of appendicitis were delayed during the COVID period with a median duration of symptoms prior to hospital attendance of two days versus one day ( =0.003) with individuals presenting with higher median white cell count than during the control period (14.9 vs 13.3, =0.031). Use of preoperative CT scanning (OR 3.013, 95% CI 1.694-5.358, <0.001) increased significantly. More complex appendicectomies (AAST grade >1) were performed (OR 2.102, 95% CI 1.155-3.826, =0.015) with a greater consultant presence during operations (OR 4.740, 95% CI 2.523-8.903, <0.001). Despite the greater AAST scores recorded during the COVID period, no increase in postoperative complications was observed (OR 1.145, 95% CI 0.404-3.244, =0.798). Delayed presentations during the COVID-19 pandemic were associated with more complex cases of appendicitis. Important lessons can be learnt from the changes in practice employed as a result of this global pandemic.

Background. Nevirapine (NVP) resistance emerges in up to 70% of women exposed to single-dose (sd) NVP for prevention of mother-to-child transmission of human immunodeficiency virus (HIV). Methods. HIV-infected pregnant women were randomized to receive sdNVP and either zidovudine/lamivudine (3TC), tenofovir/emtricitabine (FTC), or lopinavir/ritonavir for either 7 or 21 days. The primary endpoint was the emergence of new NVP resistance mutations as detected by standard population genotype at 2 and 6 weeks after treatment. Low-frequency NVP- or 3TC/FTC-resistant mutants at codons 103, 181, and 184 were sought using allele-specific polymerase chain reaction (ASP). Results. Among 484 women randomized, 422 (87%) received study treatment. Four hundred twelve (98%) women had primary endpoint results available; of these, 5 (1.2%) had new NVP resistance detected by population genotype: 4 of 215 in the 7-day arms (1.9%; K103N in 4 women with Y181C, Y188C, or G190A in 3 of 4) and 1 of 197 (0.5%; V108I) in the 21-day arms (P = .37). Among women with ASP results, new NVP resistance mutations emerged significantly more often in the 7-day arms (13/74 [18%]) than in the 21-day arms (3/66 [5%], P = .019). 3TC/FTC-resistant mutants (M184V/I) emerged infrequently (7/134 [5%]), and their occurrence did not differ by arm. Conclusions. Three short-term antiretroviral strategies, begun simultaneously with the administration of sdNVP, resulted in a low rate (1.2%) of new NVP-resistance mutations when assessed at 2 and 6 weeks following completion of study treatment by standard genotype. ASP revealed that 21-day regimens were significantly better than 7-day regimens at preventing the emergence of minor NVP resistance variants. Clinical Trials Registration. NCT00099632.

Objective. Nevirapine is metabolized by cytochrome P450 (CYP) 2B6 and CYP3A4. We characterized relationships between clinical parameters, human genetics, pharmacokinetics, and human immunodeficiency virus type 1 (HIV-1) drug resistance mutations in pregnant women following single-dose intrapartum nevirapine. Methods. In AIDS Clinical Trials Group study A5207, women received nevirapine at onset of labor and were randomly assigned to receive lamivudine/zidovudine, emtricitabine/tenofovir, or lopinavir/ritonavir for 7 or 21 days. Plasma nevirapine level was quantified on postpartum day 1 and on weeks 1, 3, and 5. We assayed 214 polymorphisms in CYP2B6 and other genes and evaluated associations with pharmacokinetic parameters, including elimination constant, time to protein-adjusted 50% inhibitory concentration (IC₅₀), and week 5 nevirapine level below the quantification limit. Results. Among 301 women with evaluable pharmacokinetic and genotype data, lower body mass index and random assignment to receive lopinavir/ritonavir were associated with more rapid nevirapine elimination. Among those of African ancestry, longer time to IC₅₀ was associated with CYP2B6 983T→C (P = .004) but not with CYP2B6 516G→T (P = .8). Among Indians, slower nevirapine elimination was associated with CYP2B6 516G→T (P = .04). Emergent resistance was infrequent and not associated with pharmacokinetics or CYP2B6 genotype. Conclusions. The effects on plasma drug exposure following single-dose nevirapine may be greater for CYP2B6 983T→C than for 516G→T and are less pronounced than at steady state.