Mitotic Exit: Thresholds and Targets

Cyclin dependent kinases (CDKs) are at the heart of the cell cycle. Throughout the cycle, these complexes modify many proteins, changing various aspects of their regulation (stability, localization, etc.). As cells exit mitosis, the CDK that has driven many of the cell cycle processes is inhibited and degraded, allowing many of the kinase substrates to return to their unphosphorylated state. This assures that each subsequent cell cycle is begun in the same naïve state, again ready for CDK-dependent regulation. The studies in this thesis focus on two mechanisms by which this restoration is accomplished in the budding yeast, Saccharomyces cerevisiae: (1) a transcriptional program that transcribes many of the genes required for physically dividing the mother and daughter cells and beginning the next round of cell division and (2) a phosphatase that specifically removes the phosphates from sites modified by CDK during exit from mitosis. Two transcription factors, Swi5 and Ace2, transcribe many of the genes required for physically dividing the mother and daughter cells and beginning the next round of cell division. Previously our lab has shown that locking mitotic cyclin levels, by inducing transcription of an undegradable form of the protein, causes dose-dependent delays in different cell cycle events. The first chapter addresses the contribution of the transcriptional program to this phenomenon. Interestingly, in these cells where mitotic cyclin levels were sustained, deletion of the transcription factor Swi5 increases the mitotic cyclin inhibition, specifically as it relates to budding and cytokinesis. Importantly, when phosphorylated by CDK, Swi5 is excluded from the nucleus, so in the second chapter, we investigate its localization when mitotic cyclin levels are locked. Swi5 still enters the nucleus. In fact in some cells, Swi5 enters the nucleus several times before the cell cycle advances. Given previous studies from our lab showing that the release of Cdc14 phosphatase also oscillates under these conditions, the reentry of Swi5 may support a model that a kinase/phosphatase balance allows cell cycle progression in these cells. All this suggests that Swi5 promotes the transcription of genes important for promoting cytokinesis and budding despite high mitotic cyclin levels. In the third chapter, we begin to assess the contribution of specific targets of the mitotic exit transcriptional program to the mitotic cyclin-dependent regulation of specific cell cycle events. Finally, Cdc14, a phosphatase that removes the phosphate groups added by CDKs, is sequestered for most of the cell cycle but released from the nucleolus during the end of mitosis. In the fourth chapter, we examine the physiological relevance of these dephosphorylation events on novel targets of the Cdc14 phosphatase

Mammal responses to global changes in human activity vary by trophic group and landscape

Wildlife must adapt to human presence to survive in the Anthropocene, so it is critical to understand species responses to humans in different contexts. We used camera trapping as a lens to view mammal responses to changes in human activity during the COVID-19 pandemic. Across 163 species sampled in 102 projects around the world, changes in the amount and timing of animal activity varied widely. Under higher human activity, mammals were less active in undeveloped areas but unexpectedly more active in developed areas while exhibiting greater nocturnality. Carnivores were most sensitive, showing the strongest decreases in activity and greatest increases in nocturnality. Wildlife managers must consider how habituation and uneven sensitivity across species may cause fundamental differences in human–wildlife interactions along gradients of human influence.Peer reviewe

Whole Exome Sequencing Analysis of Individuals with Autism Spectrum Disorder

Autism Spectrum Disorder (ASD) is a neurodevelopmental disorder characterized by deficits in social communication and interaction and the presence of repetitive behaviors and or restricted interests. ASD has heterogeneous genetic and environmental etiologies, but for most individuals with ASD the genetic cause is unknown. Determining the genetic factors contributing to an individual’s ASD phenotype is a goal in the clinic setting, as well as an important mechanism for understanding the broader pathology of ASD. As part of a larger research study, individuals with ASD and some family members had whole exome sequencing. For six probands with samples from both parents available, potential de novo variants were identified and assessed for their ability to contribute to the proband’s ASD phenotype. Some variants were in genes previously associated with non-syndromic ASD, and variants in common pathways across different probands were identified, including those involved in axon guidance. For an additional proband an analysis of the chromosomal region identified as a small deletion by a clinical microarray was completed to assess if sequence level information could contribute to the interpretation of this result. The analysis lessened the suspicion that this copy number variant contributed to the proband’s ASD phenotype. Future analysis of this data set, especially inherited variants, will continue to identify biological processes that affect the complex ASD phenotype. As an emerging technology in the genetics clinic and as a research tool, whole exome sequencing will contribute to our understanding of ASD and identify genetic targets for future discovery, diagnostics, and therapeutics

Analysis of the mitotic exit control system using locked levels of stable mitotic cyclin

Cyclin-dependent kinase (Cdk) both promotes mitotic entry (spindle assembly and anaphase) and inhibits mitotic exit (spindle disassembly and cytokinesis), leading to an elegant quantitative hypothesis that a single cyclin oscillation can function as a ratchet to order these events. This ratchet is at the core of a published ODE model for the yeast cell cycle. However, the ratchet model requires appropriate cyclin dose–response thresholds. Here, we test the inhibition of mitotic exit in budding yeast using graded levels of stable mitotic cyclin (Clb2). In opposition to the ratchet model, stable levels of Clb2 introduced dose-dependent delays, rather than hard thresholds, that varied by mitotic exit event. The ensuing cell cycle was highly abnormal, suggesting a novel reason for cyclin degradation. Cdc14 phosphatase antagonizes Clb2–Cdk, and Cdc14 is released from inhibitory nucleolar sequestration independently of stable Clb2. Thus, Cdc14/Clb2 balance may be the appropriate variable for mitotic regulation. Although our results are inconsistent with the aforementioned ODE model, revision of the model to allow Cdc14/Clb2 balance to control mitotic exit corrects these discrepancies, providing theoretical support for our conclusions

Precision Medicine Screening Using Whole Genome Sequencing and Advanced Imaging To Identify Disease Risk in Adults

ABSTRACTBACKGROUNDProgress in science and technology have created the capabilities and alternatives to symptom-driven medical care. Reducing premature mortality associated with age-related chronic diseases, such as cancer and cardiovascular disease, is an urgent priority we address using advanced screening detection.METHODSWe enrolled active adults for early detection of risk for age-related chronic disease associated with premature mortality. Whole genome sequencing together with: global metabolomics, 3D/4D imaging using non-contrast whole body magnetic resonance imaging and echocardiography, and 2-week cardiac monitoring were employed to detect age-related chronic diseases and risk for diseases.RESULTSWe detected previously unrecognized age-related chronic diseases requiring prompt (&lt;30 days) medical attention in 17 (8%, 1:12) of 209 study participants, including 4 participants with early stage neoplasms (2%, 1:50). Likely mechanistic genomic findings correlating with clinical data were identified in 52 participants (25%. 1:4). More than three-quarters of participants (n=164, 78%, 3:4) had evidence of age-related chronic diseases or associated risk factors.CONCLUSIONSPrecision medicine screening using genomics with other advanced clinical data among active adults identified unsuspected disease risks for age-related chronic diseases associated with premature mortality. This technology-driven phenotype screening approach has the potential to extend healthy life among active adults through improved early detection and prevention of age-related chronic diseases. Our success provides a scalable strategy to move medical practice and discovery toward risk detection and disease modification thus achieving healthier extension of life.SIGNIFICANCE STATEMENTAdvances in science and technology have enabled scientists to analyze the human genome cost-effectively and to combine genome sequencing with noninvasive imaging technologies for alternatives to symptom-driven medical care. Using whole genome sequencing and noninvasive 3D/4D imaging technologies we screened 209 adults to detect age-related chronic diseases, such as cancer and cardiovascular disease. We found unrecognized age-related chronic diseases requiring prompt (&lt;30 days) medical attention in 1:12 study participants, likely genomic findings correlating with clinical data in 1:4 participants, and evidence of age-related chronic diseases or associated risk factors in more than 3 of 4 participants. These results demonstrate that genome sequencing with clinical imaging data can be used for screening and early detection of diseases associated with premature mortality.</jats:sec

Precision medicine screening using whole-genome sequencing and advanced imaging to identify disease risk in adults.

Reducing premature mortality associated with age-related chronic diseases, such as cancer and cardiovascular disease, is an urgent priority. We report early results using genomics in combination with advanced imaging and other clinical testing to proactively screen for age-related chronic disease risk among adults. We enrolled active, symptom-free adults in a study of screening for age-related chronic diseases associated with premature mortality. In addition to personal and family medical history and other clinical testing, we obtained whole-genome sequencing (WGS), noncontrast whole-body MRI, dual-energy X-ray absorptiometry (DXA), global metabolomics, a new blood test for prediabetes (Quantose IR), echocardiography (ECHO), ECG, and cardiac rhythm monitoring to identify age-related chronic disease risks. Precision medicine screening using WGS and advanced imaging along with other testing among active, symptom-free adults identified a broad set of complementary age-related chronic disease risks associated with premature mortality and strengthened WGS variant interpretation. This and other similarly designed screening approaches anchored by WGS and advanced imaging may have the potential to extend healthy life among active adults through improved prevention and early detection of age-related chronic diseases (and their risk factors) associated with premature mortality

Hemoglobin-derived Peptides as Novel Type of Bioactive Signaling Molecules

Most bioactive peptides are generated by proteolytic cleavage of large precursor proteins followed by storage in secretory vesicles from where they are released upon cell stimulation. Examples of such bioactive peptides include peptide neurotransmitters, classical neuropeptides, and peptide hormones. In the last decade, it has become apparent that the breakdown of cytosolic proteins can generate peptides that have biological activity. A case in point and the focus of this review are hemoglobin-derived peptides. In vertebrates, hemoglobin (Hb) consists of a tetramer of two α- and two β-globin chains each containing a prosthetic heme group, and is primarily involved in oxygen delivery to tissues and in redox reactions (Schechter Blood 112:3927–3938, 2008). The presence of α- and/or β-globin chain in tissues besides red blood cells including rodent and human brain and peripheral tissues (Liu et al. Proc Natl Acad Sci USA 96:6643–6647, 1999; Newton et al. J Biol Chem 281:5668–5676, 2006; Wride et al. Mol Vis 9:360–396, 2003; Setton-Avruj Exp Neurol 203:568–578, 2007; Ohyagi et al. Brain Res 635:323–327, 1994; Schelshorn et al. J Cereb Blood Flow Metab 29:585–595, 2009; Richter et al. J Comp Neurol 515:538–547, 2009) suggests that globins and/or derived peptidic fragments might play additional physiological functions in different tissues. In support of this hypothesis, a number of Hb-derived peptides have been identified and shown to have diverse functions (Ivanov et al. Biopoly 43:171–188, 1997; Karelin et al. Neurochem Res 24:1117–1124, 1999). Modern mass spectrometric analyses have helped in the identification of additional Hb peptides (Newton et al. J Biol Chem 281:5668–5676, 2006; Setton-Avruj Exp Neurol 203:568–578, 2007; Gomes et al. FASEB J 23:3020–3029, 2009); the molecular targets for these are only recently beginning to be revealed. Here, we review the status of the Hb peptide field and highlight recent reports on the identification of a molecular target for a novel set of Hb peptides, hemopressins, and the implication of these peptides to normal cell function and disease. The potential therapeutic applications for these Hb-derived hemopressin peptides will also be discussed

Mammal responses to global changes in human activity vary by trophic group and landscape

AbstractWildlife must adapt to human presence to survive in the Anthropocene, so it is critical to understand species responses to humans in different contexts. We used camera trapping as a lens to view mammal responses to changes in human activity during the COVID-19 pandemic. Across 163 species sampled in 102 projects around the world, changes in the amount and timing of animal activity varied widely. Under higher human activity, mammals were less active in undeveloped areas but unexpectedly more active in developed areas while exhibiting greater nocturnality. Carnivores were most sensitive, showing the strongest decreases in activity and greatest increases in nocturnality. Wildlife managers must consider how habituation and uneven sensitivity across species may cause fundamental differences in human–wildlife interactions along gradients of human influence