Altered nuclear distribution of recombination protein RAD51 in maize mutants suggests the involvement of RAD51 in meiotic homology recognition

The recombination protein RAD51 is a component of the meiotic recombination pathway and has been proposed to play a role in the homology search, a process by which homologous chromosomes find each other before they pair in the prophase of meiosis. To study the relationship between recombination and chromosome pairing, we examined the distribution of RAD51 foci on meiotic chromosomes in maize mutants with defects in chromosome pairing. The patterns of RAD51 distribution showed dramatic variation among the meiotic mutants. The mutants generally exhibited significant decreases in the number of RAD51 foci at zygotene, corresponding to the degree of their pairing defects. These results provide evidence for a key role of RAD51 structures in the homology search.     

Cdk1/cyclin B1 controls Fas-mediated apoptosis by regulating caspase-8 activity

Caspase activation is a hallmark of apoptosis. However, the molecular mechanisms underlying the regulation of caspase-8 activation within the extrinsic death pathway are not well understood. In this study, we demonstrate that procaspase-8 is phosphorylated in mitotic cells by Cdk1/cyclin B1 on Ser-387, which is located at the N terminus of the catalytic subunit p10. This phosphorylation of procaspase-8 on Ser-387 occurs in cancer cell lines, as well as in primary breast tissues and lymphocytes. Furthermore, RNA interference-mediated silencing of cyclin B1 or treatment with the Cdk1 inhibitor RO-3306 enhances the Fas-mediated activation and processing of procaspase-8 in mitotic cells. A nonphosphorylatable procaspase-8 (S387A) facilitates Fas-induced apoptosis during mitosis. Our findings suggest that Cdk1/cyclin B1 activity shields human cells against extrinsic death stimuli and unravel the molecular details of the cross talk between cell cycle and extrinsic apoptotic pathways. Finally, this new mechanism may also contribute to tumorigenesis.     

Pivotal Role of Mitochondrial Calcium Uptake in Neural Cell Apoptosis and Necrosis

Abstract : Perturbed cellular calcium homeostasis has been implicated in both apoptosis and necrosis, but the role of altered mitochondrial calcium handling in the cell death process is unclear. The temporal ordering of changes in cytoplasmic ([Ca²⁺]C) and intramitochondrial ([Ca²⁺]M) calcium levels in relation to mitochondrial reactive oxygen species (ROS) accumulation and membrane depolarization (MD) was examined in cultured neural cells exposed to either an apoptotic (staurosporine ; STS) or a necrotic (the toxic aldehyde 4-hydroxynonenal ; HNE) insult. STS and HNE each induced an early increase of [Ca²⁺]C followed by delayed increase of [Ca²⁺]M. Overexpression of Bcl-2 blocked the elevation of [Ca²⁺]M and the MD in cells exposed to STS but not in cells exposed to HNE. The cytoplasmic calcium chelator BAPTA-AM and the inhibitor of mitochondrial calcium uptake ruthenium red prevented both apoptosis and necrosis. STS and HNE each induced mitochondrial ROS accumulation and MD, which followed the increase of [Ca²⁺]M. Cyclosporin A prevented both apoptosis and necrosis, indicating critical roles for MD in both forms of cell death. Caspase activation occurred only in cells undergoing apoptosis and preceded increased [Ca²⁺]M. Collectively, these findings suggest that mitochondrial calcium overload is a critical event in both apoptotic and necrotic cell death.     

The Impact of Immunosenescence on Humoral Immune Response Variation after Influenza A/H1N1 Vaccination in Older Subjects

Background

Although influenza causes significant morbidity and mortality in the elderly, the factors underlying the reduced vaccine immunogenicity and efficacy in this age group are not completely understood. Age and immunosenescence factors, and their impact on humoral immunity after influenza vaccination, are of growing interest for the development of better vaccines for the elderly.

Methods

We assessed associations between age and immunosenescence markers (T cell receptor rearrangement excision circles – TREC content, peripheral white blood cell telomerase – TERT expression and CD28 expression on T cells) and influenza A/H1N1 vaccine-induced measures of humoral immunity in 106 older subjects at baseline and three timepoints post-vaccination.

Results

TERT activity (TERT mRNA expression) was significantly positively correlated with the observed increase in the influenza-specific memory B cell ELISPOT response at Day 28 compared to baseline (p-value=0.025). TREC levels were positively correlated with the baseline and early (Day 3) influenza A/H1N1-specific memory B cell ELISPOT response (p-value=0.042 and p-value=0.035, respectively). The expression and/or expression change of CD28 on CD4+ and/or CD8+ T cells at baseline and Day 3 was positively correlated with the influenza A/H1N1-specific memory B cell ELISPOT response at baseline, Day 28 and Day 75 post-vaccination. In a multivariable analysis, the peak antibody response (HAI and/or VNA at Day 28) was negatively associated with age, the percentage of CD8+CD28low T cells, IgD+CD27- naïve B cells, and percentage overall CD20- B cells and plasmablasts, measured at Day 3 post-vaccination. The early change in influenza-specific memory B cell ELISPOT response was positively correlated with the observed increase in influenza A/H1N1-specific HAI antibodies at Day 28 and Day 75 relative to baseline (p-value=0.007 and p-value=0.005, respectively).

Conclusion

Our data suggest that influenza-specific humoral immunity is significantly influenced by age, and that specific markers of immunosenescence (e.g., the baseline/early expression of CD28 on CD4+ and/or CD8+ T cells and T cell immune abnormalities) are correlated with different humoral immune response outcomes observed after vaccination in older individuals, and thus can be potentially used to predict vaccine immunogenicity.     

Comparison of genomes of eight species of sections <Emphasis Type="Italic">Linum</Emphasis> and <Emphasis Type="Italic">Adenolinum</Emphasis> from the genus<Emphasis Type="Italic"> Linum</Emphasis> based on chromosome banding,molecular markers and RAPD analysis

Karyotypes of species sects. Linum and Adenolinum have been studied using C/DAPI-banding, Ag-NOR staining, FISH with 5S and 26S rDNA and RAPD analysis. C/DAPI-banding patterns enabled identification of all homologous chromosome pairs in the studied karyotypes. The revealed high similarity between species L. grandiflorum (2n = 16) and L. decumbens by chromosome and molecular markers proved their close genome relationship and identified the chromosome number in L. decumbens as 2n = 16. The similarity found for C/DAPI-banding patterns between species with the same chromosome numbers corresponds with the results obtained by RAPD-analysis, showing clusterization of 16-, 18- and 30-chromosome species into three separate groups. 5S rDNA and 26S rDNA were co-localized in NOR-chromosome 1 in the genomes of all species investigated. In 30-chromosome species, there were three separate 5S rDNA sites in chromosomes 3, 8 and 13. In 16-chromosome species, a separate 5S rDNA site was also located in chromosome 3, whereas in 18-chromosome species it was found in the long arm of NOR-chromosome 1. Thus, the difference in localization of rDNA sites in species with 2n = 16, 2n = 30 and 2n = 18 confirms taxonomists opinion, who attributed these species to different sects. Linum and Adenolinum, respectively. The obtained results suggest that species with 2n = 16, 2n = 18 and 2n = 30 originated from a 16-chromosome ancestor.     

Serial cross-sectional estimation of vaccine-and infection-induced SARS-CoV-2 seroprevalence in British Columbia,Canada

Background:The evolving proportion of the population considered immunologically naive versus primed for more efficient immune memory response to SARS-CoV-2 has implications for risk assessment. We sought to chronicle vaccine- and infection-induced seroprevalence across the first 7 waves of the COVID-19 pandemic in British Columbia, Canada.Methods:During 8 cross-sectional serosurveys conducted between March 2020 and August 2022, we obtained anonymized residual sera from children and adults who attended an outpatient laboratory network in the Lower Mainland (Greater Vancouver and Fraser Valley). We used at least 3 immunoassays per serosurvey to detect SARS-CoV-2 spike and nucleocapsid antibodies. We assessed any seroprevalence (vaccineor infection-induced, or both), defined by positivity on any 2 assays, and infection-induced seroprevalence, also defined by dual-assay positivity but requiring both antinucleocapsid and antispike detection. We used estimates of infection-induced seroprevalence to explore underascertainment of infections by surveillance case reports.Results:By January 2021, we estimated that any seroprevalence remained less than 5%, increasing with vaccine rollout to 56% by May–June 2021, 83% by September–October 2021 and 95% by March 2022. Infection-induced seroprevalence remained less than 15% through September–October 2021, increasing across Omicron waves to 42% by March 2022 and 61% by July–August 2022. By August 2022, 70%–80% of children younger than 20 years and 60%–70% of adults aged 20–59 years had been infected, but fewer than half of adults aged 60 years and older had been infected. Compared with estimates of infection-induced seroprevalence, surveillance case reports underestimated infections 12-fold between September 2021 and March 2022 and 92-fold between March 2022 and August 2022.Interpretation:By August 2022, most children and adults younger than 60 years had evidence of both SARS-CoV-2 vaccination and infection. As previous evidence suggests that a history of both exposures may induce stronger, more durable hybrid immunity than either exposure alone, older adults — who have the lowest infection rates but highest risk of severe outcomes — continue to warrant prioritized vaccination.

The British Columbia Centre for Disease Control (BCCDC) has a long-established serosurvey protocol to monitor population susceptibility to emerging or re-emerging respiratory viruses. The approach was first deployed during the influenza A (H1N1) pandemic in 2009 to monitor changes in seroprevalence across successive pandemic waves and the mass vaccination campaign.^1–7 The methodology is predicated upon serial cross-sectional convenience sampling of anonymized residual sera from children and adults of all ages in the most populated Lower Mainland region of BC.^8,9Adapting this protocol, the BCCDC launched its first SARS-CoV-2 serosurvey in March 2020, just before the World Health Organization’s declaration of the COVID-19 pandemic. ¹⁰ Baseline assessment was followed by additional serosurveys that spanned the time from mRNA vaccine availability in mid-December 2020, through 7 pandemic waves associated with multiple variants of concern to August 2022 (Figure 1).^11–13 Using these serosurveys, we sought to track the evolving proportion of the population that remained immunologically naive and, thus, fully susceptible to COVID-19, versus the evolving proportion that was immunologically primed (through vaccination or infection) and, thus, poised for more efficient memory response in mitigating the risk of SARS-CoV-2. Recognizing the spectrum of illness, including asymptomatic or mild infections, and variable diagnostic access, case identification and reporting, we also used estimates of infection-induced seroprevalence to explore the potential underascertainment of infections by surveillance case reports.Open in a separate windowFigure 1:Provincial surveillance case reports to the British Columbia Centre for Disease Control (BCCDC) by epidemiological week from January 2020 to September 2022, with timing of serosurveys and select public health measures, in BC, Canada. We group case tallies by epidemiological week (7-d period) as per standard surveillance methods for comparing data by period from year to year. Epidemic waves are enumerated sequentially and are displayed with the predominant variant of concern (VOC). Publicly funded access to nucleic acid amplification tests (NAATs) or rapid antigen tests (RATs) is displayed below the X-axis. For details on public health measures, vaccines, schedules and coverage estimates, see Appendix 1, Supplementary Material 1, available at www.cmaj.ca/lookup/doi/10.1503/cmaj.221335/tab-related-content. *Nonessential travel discouraged, health care service delivery adjusted, public gatherings > 50 people prohibited. Provincial state of emergency declared. †Interactions limited to households or “core bubble” (immediate family or those in same dwelling) or to a maximum of 2 other people if living alone. ‡Dine-in food services and indoor fitness activities banned, only essential travel permitted. §Gradual return to gatherings, recreational travel, in-person work, which was interrupted by the fourth wave. ¶Indoor and personal gatherings limited, 50% capacity limit at venues of > 1000 people, sports tournaments paused. Social restrictions lifted during epidemiological week 7, 2022. **Mask mandates lifted during epidemiological week 10, 2022. ††The first 2 spike-based mRNA vaccine formulations were authorized during epidemiological weeks 50 and 52, 2020, respectively, with mRNA vaccines comprising most doses (> 90%) administered in BC and Canada across the pandemic. In epidemiological week 8, 2021, a chimpanzee adenoviral-vectored (ChAdOx1) vaccine was also authorized. ‡‡Vaccines (mRNA) initially deployed to high-risk individuals, including residents and staff of long-term care and assisted-living facilities, essential visitors within those settings and health care workers. §§Community-based vaccine roll-out, prioritized by age, beginning with the oldest adults in mid-March 2021. Access to booster doses followed similar prioritization sequence, inclusive of clinically extremely vulnerable individuals of any age. ¶¶Single-dose vaccine card required for entry into social and recreational settings starting in epidemiological week 37, 2021; 2-dose cards were required beginning in epidemiological week 43, 2021. Vaccine cards were ultimately repealed in epidemiological week 14, 2022.     

Decreased rates of cerebral protein synthesis measured in vivo in a mouse model of Tuberous Sclerosis Complex: unexpected consequences of reduced tuberin

Tuberous sclerosis complex (TSC ) is an autosomal dominant neurogenetic disorder affecting about 1 in 6000 people and is caused by mutations in either TSC 1 or TSC 2 . This disorder is characterized by increased activity of mammalian target of rapamycin complex 1 (mTORC 1), which is involved in regulating ribosomal biogenesis and translation initiation. We measured the effects of Tsc2 haploinsufficiency (Tsc2 ^+/? ) in 3‐month‐old male mice on regional rates of cerebral protein synthesis (rCPS ) by means of the in vivo L‐[1‐¹⁴C]leucine method. This quantitative autoradiographic method includes an estimate of the integrated specific activity of the tracer amino acid in brain tissue. The estimate accounts for recycling of unlabeled amino acids from tissue protein breakdown by means of a factor (λ) that was determined in control and Tsc2 ^+/? mice. The value of λ was higher in Tsc2 ^+/? mice, indicating that a greater fraction of leucine in the tissue precursor pool for protein synthesis is derived from the plasma compared to controls, consistent with reduced rates of protein degradation. We determined rCPS in freely moving, awake male Tsc2 ^+/? and control mice, and we used the determined values of λ in the calculation of rCPS . Unexpectedly, we found that rCPS were significantly decreased in 16 of the 17 brain regions analyzed in Tsc2 ^+/? mice compared to controls. Our results indicate a complex role of mTORC 1 in the regulation of cerebral protein synthesis that has not been previously recognized.

     

Genetic polymorphisms associated with rubella virus-specific cellular immunity following MMR vaccination

Rubella virus causes a relatively benign disease in most cases, although infection during pregnancy can result in serious birth defects. An effective vaccine has been available since the early 1970s and outbreaks typically do not occur among highly vaccinated (≥2 doses) populations. Nevertheless, considerable inter-individual variation in immune response to rubella immunization does exist, with single-dose seroconversion rates ~95 %. Understanding the mechanisms behind this variability may provide important insights into rubella immunity. In the current study, we examined associations between single nucleotide polymorphisms (SNPs) in selected cytokine, cytokine receptor, and innate/antiviral genes and immune responses following rubella vaccination in order to understand genetic influences on vaccine response. Our approach consisted of a discovery cohort of 887 subjects aged 11–22 at the time of enrollment and a replication cohort of 542 older adolescents and young adults (age 18–40). Our data indicate that SNPs near the butyrophilin genes (BTN3A3/BTN2A1) and cytokine receptors (IL10RB/IFNAR1) are associated with variations in IFNγ secretion and that multiple SNPs in the PVR gene, as well as SNPs located in the ADAR gene, exhibit significant associations with rubella virus-specific IL-6 secretion. This information may be useful, not only in furthering our understanding immune responses to rubella vaccine, but also in identifying key pathways for targeted adjuvant use to boost immunity in those with weak or absent immunity following vaccination.     

Backbone Cyclization of the C-terminal Part of Substance P. Part 1: The Important Role of the Sulphur in Position 11

Novel backbone-to-side chain and backbone-to-backbone cyclic analogues of substance P (SP) were prepared by solid-phase synthesis and screened for biological activity. An analogue containing a thioether- lactam ring between positions 9 and 11 showed an EC₅₀ value of 20nM toward the neurokinin 1 (NK-1) and was inactive toward the NK-2 and NK-3 receptors. On the other hand, in a multiple backbone cyclic peptide library of similar analogues, in which the sulphur was excluded from the ring, very low activity was detected. The activity was re-evaluated and was found to be even lower (EC₅₀=0.11 mM ) than the previously published data. These results indicate that the thioether moiety has a crucial role in receptor activation. The results also show tolerance of the NK-1 receptor, but not NK-2 or NK-3, to cyclization of the C-terminal portion of the SP_6–11 hexapeptide.