JIP4 is a PLK1 binding protein that regulates p38MAPK activity in G2 phase

Cell cycle progression from G2 phase into mitosis is regulated by a complex network of mechanisms, all of which finally control the timing of Cyclin B/CDK1 activation. PLK1 regulates a network of events that contribute to regulating G2/M phase progression. Here we have used a proteomics approach to identify proteins that specifically bind to the Polobox domain of PLK1. This identified a panel of proteins that were either associated with PLK1 in G2 phase and/or mitosis, the strongest interaction being with the MAPK scaffold protein JIP4. PLK1 binding to JIP4 was found in G2 phase and mitosis, and PLK1 binding was self-primed by PLK1 phosphorylation of JIP4. PLK1 binding is required for JIP4-dependent p38MAPK activation in G2 phase during normal cell cycle progression, but not in either G2 phase or mitotic stress response. Finally, JIP4 is a target for caspase-dependent cleavage in mitotically arrested cells. The role for the PLK1–JIP4 regulated p38MAPK activation in G2 phase is unclear, but it does not affect either progression into or through mitosis

Establishment of a gnotobiotic pig model of Clostridioides difficile infection and disease

Clostridioides difficile (C. difficile) is a gram-positive, spore-forming, anaerobic bacterium known to be the most common cause of hospital-acquired and antibiotic-associated diarrhea. C. difficile infection rates are on the rise worldwide and treatment options are limited, indicating a clear need for novel therapeutics. Gnotobiotic piglets are an excellent model to reproduce the acute pseudomembranous colitis (PMC) caused by C. difficile due to their physiological similarities to humans and high susceptibility to infection. Here, we established a gnotobiotic pig model of C. difficile infection and disease using a hypervirulent strain. C. difficile-infected pigs displayed classic signs of C. difficile infection, including severe diarrhea and weight loss. Inoculated pigs had severe gross and microscopic intestinal lesions. C. difficile infection caused an increase in pro-inflammatory cytokines in samples of serum, large intestinal contents, and pleural effusion. C. difficile spores and toxins were detected in the feces of inoculated animals as tested by anaerobic culture and cytotoxicity assays. Successful establishment of this model is key for future work as therapeutics can be evaluated in an environment that accurately mimics what happens in humans. The model is especially suitable for evaluating potential prophylactics and therapeutics, including vaccines and passive immune strategies.Instituto de VirologíaFil: Nyblade, Charlotte. Virginia Polytechnic Institute and State University. Virginia-Maryland College of Veterinary Medicine. Department of Biomedical Sciences and Pathobiology; Estados UnidosFil: Parreño, Gladys Viviana. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Virología e Innovaciones Tecnológicas; ArgentinaFil: Parreño, Gladys Viviana. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Parreño, Gladys Viviana. Virginia Polytechnic Institute and State University. Virginia-Maryland College of Veterinary Medicine. Department of Biomedical Sciences and Pathobiology; Estados UnidosFil: Zhou, Peng. Virginia Polytechnic Institute and State University. Virginia-Maryland College of Veterinary Medicine. Department of Biomedical Sciences and Pathobiology; Estados UnidosFil: Hensley, Casey. Virginia Polytechnic Institute and State University. Virginia-Maryland College of Veterinary Medicine. Department of Biomedical Sciences and Pathobiology; Estados UnidosFil: Oakes, Vanessa. Virginia Polytechnic Institute and State University. Virginia-Maryland College of Veterinary Medicine. Department of Biomedical Sciences and Pathobiology; Estados UnidosFil: Mahsoub, Hassan M. Virginia Polytechnic Institute and State University. Virginia-Maryland College of Veterinary Medicine. Department of Biomedical Sciences and Pathobiology; Estados UnidosFil: Mahsoub, Hassan M. Virginia Polytechnic Institute and State University. Center for Emerging, Zoonotic, and Arthropod‑Borne Pathogens; Estados UnidosFil: Kiley, Kelsey. Virginia Polytechnic Institute and State University. Virginia-Maryland College of Veterinary Medicine. Department of Biomedical Sciences and Pathobiology; Estados UnidosFil: Frazier, Maggie. Virginia Polytechnic Institute and State University. Virginia-Maryland College of Veterinary Medicine. Department of Biomedical Sciences and Pathobiology; Estados UnidosFil: Frazier, Annie. Virginia Polytechnic Institute and State University. Virginia-Maryland College of Veterinary Medicine. Department of Biomedical Sciences and Pathobiology; Estados UnidosFil: Zhang, Yongrong. University of Maryland at Baltimore. Department of Microbial Pathogenesis; Estados UnidosFil: Feng, Hanping. University of Maryland at Baltimore. Department of Microbial Pathogenesis; Estados UnidosFil: Yuan, Lijuan. Virginia Polytechnic Institute and State University. Virginia-Maryland College of Veterinary Medicine. Department of Biomedical Sciences and Pathobiology; Estados UnidosFil: Yuan, Lijuan. Virginia Polytechnic Institute and State University. Center for Emerging, Zoonotic, and Arthropod‑Borne Pathogens; Estados Unido

CDC25B associates with a centrin 2-containing complex and is involved in maintaining centrosome integrity

Background information. CDC25 (cell division cycle 25) phosphatases function as activators of CDK (cyclin-dependent kinase)–cyclin complexes to regulate progression through the CDC. We have recently identified a pool of CDC25B at the centrosome of interphase cells that plays a role in regulating centrosome numbers

Cyclin A/CDK2 regulates multiple G2 phase pathways

The cell cycle is a carefully choreographed series of phases that when executed successfully will allow the complete replication of the genome and the equal division of the genome and other cellular content into two independent daughter cells. The inability of the cell to execute cell division successfully can result in either checkpoint activation to allow repair and/or apoptosis and/or mutations/errors that may or may not lead to tumourgenesis. Cyclin A/CDK2 is the primary cyclin/CDK regulating G2 phase progression of the cell cycle. Cyclin A/CDK2 activity peaks in G2 phase and its inhibition causes a G2 phase delay that we have termed 'the cyclin A/CDK2 dependent G2 delay'. Understanding the key pathways that are involved in the cyclin A/CDK2 dependent G2 delay has been the primary focus of this study. Characterising the cyclin A/CDK2 dependent G2 delay revealed accumulated levels of the inactive form of the mitotic regulator, cyclin B/CDK1. Surprisingly, there was also increased microtubule nucleation at the centrosomes, and the centrosomes stained for markers of cyclin B/CDK1 activity. Both microtubule nucleation at the centrosomes and phosphoprotein markers were lost with short-term treatment of CDK1/2 inhibition. Cyclin A/CDK2 localised at the centrosomes in late G2 phase after separation of the centrosomes but before the start of prophase. Thus G2 phase cyclin A/CDK2 controls the timing of entry into mitosis by controlling the subsequent activation of cyclin B/CDK1, but also has an unexpected role in coordinating the activation of cyclin B/CDK1 at the centrosome and in the nucleus. In addition to regulating the timing of cyclin B/CDK1 activation and entry into mitosis in the unperturbed cell cycle, cyclin A/CDK2 also was shown to have a role in G2 phase checkpoint recovery. Known G2 phase regulators were investigated to determine whether they had a role in imposing the cyclin A/ CDK2 dependent G2 delay. Examination of the critical G2 checkpoint arrest protein, Chk1, which also has a role during unperturbed G2/M phases revealed the presence of activated Chk1 in G2 phase, in a range of cell lines. Activated Chk1 levels were shown to accumulate in cyclin A/CDK2 depleted/inhibited cells. Further investigations revealed that Chk1, but not Chk2, depletion could reverse the cyclin A/CDK2 dependent G2 delay. It was confirmed that the accumulative activation of Chk1 was not a consequence of DNA damage induced by cyclin A depletion. The potential of cyclin A/CDK2 to regulate Chk1 revealed that the inhibitory phosphorylations, Ser286 and Ser301, were not directly catalysed by cyclin A/CDK2 in G2 phase to regulate mitotic entry. It appeared that the ability of cyclin A/CDK2 to regulate cyclin B/CDK1 activation impacted cyclin B/CDK1s phosphorylation of Chk1 on Ser286 and Ser301, thereby contributing to the delay in G2/M phase progression. Chk1 inhibition/depletion partially abrogated the cyclin A/CDK2 dependent G2 delay, and was less effective in abrogating G2 phase checkpoint suggesting that other cyclin A/CDK2 dependent mechanisms contributed to these roles of cyclin A/CDK2. In an attempt to identify these other contributing factors another G2/M phase regulator known to be regulated by cyclin A/CDK2, Cdh1 and its substrates Plk1 and Claspin were examined. Cdh1 levels were reduced in cyclin A/CDK2 depleted/inhibited cells although this had little effect on Plk1, a known Cdh1 substrate. However, the level of another substrate, Claspin, was increased. Cdh1 depletion mimicked the effect of cyclin A depletion but to a weaker extent and was sufficient at increasing Claspin levels similar to the increase caused by cyclin A depletion. Co-depletion of cyclin A and Claspin blocked the accumulation of activated Chk1 normally seen with cyclin A depletion alone. However Claspin depletion alone did not reduce the cyclin A/CDK2 dependent G2 delay but this is likely to be a result of inhibition of S phase roles of Claspin. Together, these data suggest that cyclin A/CDK2 regulates a number of different mechanisms that contribute to G2/M phase progression. Here it has been demonstrated that in normal G2/M progression and possibly to a lesser extent in G2 phase checkpoint recovery, cyclin A/CDK2 regulates the level of Cdh1 which in turn affects at least one of its substrates, Claspin, and consequently results in the increased level of activated Chk1 observed. However, the involvement of Cdh1 and Claspin alone does not explain the G2 phase delay observed with cyclin A/CDK2 depletion/inhibition. It is likely that other mechanisms, possibly including cyclin A/CDK2 regulation of Wee1 and FoxM1, as reported by others, combine with the mechanism described here to regulate normal G2/M phase progression and G2 phase checkpoint recovery. These findings support the critical role for cyclin A/CDK2 in regulating progression into mitosis and suggest that upstream regulators of cyclin A/CDK2 activation will also be critical controllers of this cell cycle transition. The pathways that work to co-ordinate cell cycle progression are very intricate and deciphering these pathways, required for normal cell cycle progression, is key to understanding tumour development. By understanding cell cycle regulatory pathways it will allow the identification of the pathway/s and their mechanism/s that become affected in tumourgenesis. This will lead to the development of better targeted therapies, inferring better efficacy with fewer side effects than commonly seen with the use of traditional therapies, such as chemotherapy. Furthermore, this has the potential to positively impact the development of personalised medicines and the customisation of healthcare

Natural Variability in Parent-Child Puzzle Play at Home

Here, we observed 3- to 4-year-old children (N=31) and their parents playing with puzzles at home during a zoom session to provide insight into the variability of the kinds of puzzles children have in their home, and the variability in how children and their parents play with spatial toys. We observed a large amount of variability in both children and parents' behaviors, and in the puzzles they selected. Further, we found relations between parents' and children's behaviors. For example, parents provided more scaffolding behaviors for younger children and parents' persistence-focused language was related to more child attempts after failure. Altogether, the present work shows how using methods of observing children at a distance, we can gain insight into the environment in which they are developing. The results are discussed in terms of how variability in spatial toys and spatial play during naturalistic interactions can help us contextualize the conclusions we draw from lab-based studies

Developing an Understanding of Emotion Categories: Lessons from Objects

How and when infants and young children begin to develop emotion categories is not yet well understood. Research has largely treated the learning problem as one of identifying perceptual similarities among exemplars (typically posed, stereotyped facial configurations). However, recent meta-analyses and reviews converge to suggest that emotion categories are abstract, involving high-dimensional and situationally variable instances. In this paper we consult research on the development of abstract object categorization to guide hypotheses about how infants might learn abstract emotion categories because the two domains present infants with similar learning challenges. In particular, we consider how a developmental cascades framework offers opportunities to understand how and when young children develop emotion categories

A Novel Approach to Assessing Infant and Child Mental Rotation.

Mental rotation is a critically important, early developing spatial skill that is related to other spatial cognitive abilities. Understanding the early development of this skill, however, requires a developmentally appropriate assessment that can be used with infants, toddlers, and young children. We present here a new eye-tracking task that uses a staircase procedure to assess mental rotation in 12-, 24-, and 36-month-old children (N = 41). To ensure that all children understood the task, the session began with training and practice, in which the children learned to fixate which of two houses a giraffe, facing either left or right, would approach. The adaptive two-up, one-down staircase procedure assessed the childrens ability to fixate the correct house when the giraffe was rotated in 30° (up) or 15° (down) increments. The procedure was successful, with most children showing evidence of mental rotation. In addition, the children were less likely to succeed as the angle of rotation increased, and the older children succeeded at higher angles of rotation than the younger children, replicating previous findings with other procedures. The present study contributes a new paradigm that can assess the development of mental rotation in young children and holds promise for yielding insights into individual differences in mental rotation

Developmental Changes in Children's Object Insertions during Play

This project contains supplemental materials for "Developmental changes in children's object insertions during play". Here you can find the demographics of the included and excluded children, sample video of children performing the play task, the R scripts used to conduct the analysis and generate the figures for the data, and the data in both processed and raw form. All files are located in subfolders under the main folder "ShapeSorterAnalysis"