49 research outputs found

    Mammalian MCM Loading in Late-G1 Coincides with Rb Hyperphosphorylation and the Transition to Post-Transcriptional Control of Progression into S-Phase

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    BACKGROUND: Control of the onset of DNA synthesis in mammalian cells requires the coordinated assembly and activation of the pre-Replication Complex. In order to understand the regulatory events controlling preRC dynamics, we have investigated how the timing of preRC assembly relates temporally to other biochemical events governing progress into S-phase. METHODOLOGY/PRINCIPAL FINDING: In murine and Chinese hamster (CHO) cells released from quiescence, the loading of the replicative MCM helicase onto chromatin occurs in the final 3-4 hrs of G(1). Cdc45 and PCNA, both of which are required for G(1)-S transit, bind to chromatin at the G(1)-S transition or even earlier in G(1), when MCMs load. An RNA polymerase II inhibitor (DRB) was added to synchronized murine keratinocytes to show that they are no longer dependent on new mRNA synthesis 3-4 hrs prior to S-phase entry, which is also true for CHO and human cells. Further, CHO cells can progress into S-phase on time, and complete S-phase, under conditions where new mRNA synthesis is significantly compromised, and such mRNA suppression causes no adverse effects on preRC dynamics prior to, or during, S-phase progression. Even more intriguing, hyperphosphorylation of Rb coincides with the start of MCM loading and, paradoxically, with the time in late-G(1) when de novo mRNA synthesis is no longer rate limiting for progression into S-phase. CONCLUSIONS/SIGNIFICANCE: MCM, Cdc45, and PCNA loading, and the subsequent transit through G(1)-S, do not depend on concurrent new mRNA synthesis. These results indicate that mammalian cells pass through a distinct transition in late-G(1) at which time Rb becomes hyperphosphorylated and MCM loading commences, but that after this transition the control of MCM, Cdc45, and PCNA loading and the onset of DNA replication are regulated at the post-transcriptional level

    Speed and Convenience Aren't Everything with Diagnostics

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    Sleep and other factors associated with mental health and psychological distress after intensive care for critical illness

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    Purpose: Some patients who survive intensive care unit (ICU) treatment report psychological sequelae during recovery. This study examined factors associated with psychological outcomes of former ICU patients up to 6 months after hospital discharge. Methods: Participants (n = 195) were adult survivors of ICU enrolled in a multicenter trial of physical rehabilitation after hospital discharge. The 36-Item Short-Form Health Survey (SF-36), the Impact of Events Scale (IES) and the Depression, Anxiety and Stress Scales were completed, and sleep rated on a five-point scale at weeks 1, 8 and 26; clinical and demographic data were obtained from patient records. Results: Participants were 41% females with mean ยฑ standard deviation (SD) age of 57 ยฑ 16 years and Acute Physiology and Chronic Health Evaluation II (APACHE II) scores of 19 ยฑ 7; median lengths of mechanical ventilation and ICU stay were 89 h and 6 days, respectively. Impaired mental health, depression, anxiety, stress and psychological distress significantly improved after week 1. Female gender, younger age and sleeping problems were associated with impaired psychological outcomes on bivariate analyses. Age; gender; week 1 anxiety, depression and stress; week 26 sleeping; and rehabilitation study group were entered into multiple linear regression analyses for week 26 IES and SF-36 Mental Component Summary (MCS) outcomes. IES scores were associated (p < 0.05) with gender, week 1 stress and week 26 sleep but not study group; MCS scores were associated (p < 0.05) with week 1 stress and week 26 sleep but not study group.Conclusion: Female gender, early levels of increased stress and problems sleeping are associated with worse psychological recovery for survivors of critical illness
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