Ecdysone signaling induces two phases of cell cycle exit in Drosophila cells

During development, cell proliferation and differentiation must be tightly coordinated to ensure proper tissue morphogenesis. Because steroid hormones are central regulators of developmental timing, understanding the links between steroid hormone signaling and cell proliferation is crucial to understanding the molecular basis of morphogenesis. Here we examined the mechanism by which the steroid hormone ecdysone regulates the cell cycle in Drosophila. We find that a cell cycle arrest induced by ecdysone in Drosophila cell culture is analogous to a G2 cell cycle arrest observed in the early pupa wing. We show that in the wing, ecdysone signaling at the larva-to-puparium transition induces Broad which in turn represses the cdc25c phosphatase String. The repression of String generates a temporary G2 arrest that synchronizes the cell cycle in the wing epithelium during early pupa wing elongation and flattening. As ecdysone levels decline after the larva-to-puparium pulse during early metamorphosis, Broad expression plummets, allowing String to become re-activated, which promotes rapid G2/M progression and a subsequent synchronized final cell cycle in the wing. In this manner, pulses of ecdysone can both synchronize the final cell cycle and promote the coordinated acquisition of terminal differentiation characteristics in the wing

Combinatorial control of temporal gene expression in the Drosophila wing by enhancers and core promoters

Abstract Background The transformation of a developing epithelium into an adult structure is a complex process, which often involves coordinated changes in cell proliferation, metabolism, adhesion, and shape. To identify genetic mechanisms that control epithelial differentiation, we analyzed the temporal patterns of gene expression during metamorphosis of the Drosophila wing. Results We found that a striking number of genes, approximately 50% of the Drosophila transcriptome, exhibited changes in expression during a time course of wing development. While cis-acting enhancer sequences clearly correlated with these changes, a stronger correlation was discovered between core-promoter types and the dynamic patterns of gene expression within this differentiating tissue. In support of the hypothesis that core-promoter type influences the dynamics of expression, expression levels of several TATA-box binding protein associated factors (TAFs) and other core promoter-associated components changed during this developmental time course, and a testes-specific TAF (tTAF) played a critical role in timing cellular differentiation within the wing. Conclusions Our results suggest that the combinatorial control of gene expression via cis-acting enhancer sequences and core-promoter types, determine the complex changes in gene expression that drive morphogenesis and terminal differentiation of the Drosophila wing epithelium.http://deepblue.lib.umich.edu/bitstream/2027.42/112935/1/12864_2012_Article_4965.pd

Developing Future Biologists: Developmental Biology for Undergraduates from Underserved Communities

Developing Future Biologists (DFB) is an inclusive, trainee-run organization that strives to excite and engage the next generation of biologists, regardless of race, gender or socioeconomic status, in the field of developmental biology. DFB offers a week-long course consisting of active lectures, hands-on laboratory sessions, and professional development opportunities through interactions with scientists from a variety of backgrounds and careers. A major goal of DFB is to propel undergraduate students from underserved communities to pursue biomedical research opportunities and advanced degrees in science. To achieve this goal, we provide DFB participants with continuing access to a diverse network of scientists that students can utilize to secure opportunities and foster success throughout multiple stages of their research careers. Here, we describe the flourishing DFB program at the University of Michigan to encourage other institutions to create their own DFB programs

Hormone-dependent control of developmental timing through regulation of chromatin accessibility

Specification of tissue identity during development requires precise coordination of gene expression in both space and time. Spatially, master regulatory transcription factors are required to control tissue-specific gene expression programs. However, the mechanisms controlling how tissue-specific gene expression changes over time are less well understood. Here, we show that hormone-induced transcription factors control temporal gene expression by regulating the accessibility of DNA regulatory elements. Using the Drosophila wing, we demonstrate that temporal changes in gene expression are accompanied by genome-wide changes in chromatin accessibility at temporal-specific enhancers. We also uncover a temporal cascade of transcription factors following a pulse of the steroid hormone ecdysone such that different times in wing development can be defined by distinct combinations of hormone-induced transcription factors. Finally, we show that the ecdysone-induced transcription factor E93 controls temporal identity by directly regulating chromatin accessibility across the genome. Notably, we found that E93 controls enhancer activity through three different modalities, including promoting accessibility of late-acting enhancers and decreasing accessibility of early-acting enhancers. Together, this work supports a model in which an extrinsic signal triggers an intrinsic transcription factor cascade that drives development forward in time through regulation of chromatin accessibility

Growth Arrest‐Specific 6 (GAS6) Promotes Prostate Cancer Survival by G1 Arrest/S Phase Delay and Inhibition of Apoptosis During Chemotherapy in Bone Marrow

Prostate cancer (PCa) is known to develop resistance to chemotherapy. Growth arrest‐specific 6 (GAS6), plays a role in tumor progression by regulating growth in many cancers. Here, we explored how GAS6 regulates the cell cycle and apoptosis of PCa cells in response to chemotherapy. We found that GAS6 is sufficient to significantly increase the fraction of cells in G1 and the duration of phase in PCa cells. Importantly, the effect of GAS6 on G1 is potentiated during docetaxel chemotherapy. GAS6 altered the levels of several key cell cycle regulators, including the downregulation of Cyclin B1 (G2/M phase), CDC25A, Cyclin E1, and CDK2 (S phase entry), while the upregulation of cell cycle inhibitors p27 and p21, Cyclin D1, and CDK4. Importantly, these changes became further accentuated during docetaxel treatment in the presence of GAS6. Moreover, GAS6 alters the apoptotic response of PCa cells during docetaxel chemotherapy. Docetaxel induced PCa cell apoptosis is efficiently suppressed in PCa cell culture in the presence of GAS6 or GAS6 secreted from co‐cultured osteoblasts. Similarly, the GAS6‐expressing bone environment protects PCa cells from apoptosis within primary tumors in vivo studies. Docetaxel induced significant levels of Caspase‐3 and PARP cleavage in PCa cells, while GAS6 protected PCa cells from docetaxel‐induced apoptotic signaling. Together, these data suggest that GAS6, expressed by osteoblasts in the bone marrow, plays a significant role in the regulation of PCa cell survival during chemotherapy, which will have important implications for targeting metastatic disease. J. Cell. Biochem. 117: 2815–2824, 2016. © 2016 Wiley Periodicals, Inc.We explored how GAS6, expressed by osteoblasts, regulates the cell cycle and apoptosis in PCa cells during chemotherapy in the bone marrow. We demonstrate that GAS6 significantly increases the number of G1 arrested cells by altering signaling networks associated with G1 arrest and S phase delay. Our results suggest that GAS6 contributes to the regulation of PCa cell survival during chemotherapy in the bone marrow microenvironment.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/134410/1/jcb25582_am.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/134410/2/jcb25582.pd

Detection and isolation of disseminated tumor cells in bone marrow of patients with clinically localized prostate cancer

BackgroundDisseminated tumor cells (DTCs) have been reported in the bone marrow (BM) of patients with localized prostate cancer (PCa). However, the existence of these cells continues to be questioned, and few methods exist for viable DTC isolation. Therefore, we sought to develop novel approaches to identify and, if detected, analyze localized PCa patient DTCs.MethodsWe used fluorescence‐activated cell sorting (FACS) to isolate a putative DTC population, which was negative for CD45, CD235a, alkaline phosphatase, and CD34, and strongly expressed EPCAM. We examined tumor cell content by bulk cell RNA sequencing (RNA‐Seq) and whole‐exome sequencing after whole genome amplification. We also enriched for BM DTCs with α‐EPCAM immunomagnetic beads and performed quantitative reverse trancriptase polymerase chain reaction (qRT‐PCR) for PCa markers.ResultsAt a threshold of 4 cells per million BM cells, the putative DTC population was present in 10 of 58 patients (17%) with localized PCa, 4 of 8 patients with metastatic PCa of varying disease control, and 1 of 8 patients with no known cancer, and was positively correlated with patients’ plasma PSA values. RNA‐Seq analysis of the putative DTC population collected from samples above (3 patients) and below (5 patients) the threshold of 4 putative DTCs per million showed increased expression of PCa marker genes in 4 of 8 patients with localized PCa, but not the one normal donor who had the putative DTC population present. Whole‐exome sequencing also showed the presence of single nucleotide polymorphisms and structural variants in the gene characteristics of PCa in 2 of 3 localized PCa patients. To examine the likely contaminating cell types, we used a myeloid colony formation assay, differential counts of cell smears, and analysis of the RNA‐Seq data using the CIBERSORT algorithm, which most strongly suggested the presence of B‐cell lineages as a contaminant. Finally, we used EPCAM enrichment and qRT‐PCR for PCa markers to estimate DTC prevalence and found evidence of DTCs in 21 of 44 samples (47%).ConclusionThese data support the presence of DTCs in the BM of a subset of patients with localized PCa and describe a novel FACS method for isolation and analysis of viable DTCs.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/151343/1/pros23896.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/151343/2/pros23896_am.pd

Ultra diffuse galaxies in the Hydra I cluster from the LEWIS Project: Phase-Space distribution and globular cluster richness

Although ultra diffuse galaxies (UDGs) are found in large numbers in clusters of galaxies, the role of the cluster environment in shaping their low surface brightness and large sizes is still uncertain. Here we examine a sample of UDGs in the Hydra I cluster (D = 51 Mpc) with new radial velocities obtained as part of the LEWIS (Looking into the faintest with MUSE) project using VLT/MUSE data. Using a phase-space, or infall diagnostic, diagram we compare the UDGs to other known galaxies in the Hydra I cluster and to UDGs in other clusters. The UDGs, along with the bulk of regular Hydra I galaxies, have low relative velocities and are located near the cluster core, and thus consistent with very early infall into the cluster. Combining with literature data, we do not find the expected trend of GC-rich UDGs associated with earlier infall times. This result suggests that quenching mechanisms other than cluster infall should be further considered, e.g. quenching by strong feedback or in cosmic sheets and filaments. Tidal stripping of GCs in the cluster environment also warrants further modelling.Comment: 6 pages, 2 figures, MNRAS, 525, 9

A robust cell cycle control mechanism limits E2F-induced proliferation of terminally differentiated cells in vivo

Overexpression of both CycE and E2F is necessary to trigger cell cycle reentry and overproliferation of terminally differentiated wing cells

Looking into the faintEst WIth MUSE (LEWIS): on the nature of ultra-diffuse galaxies in the Hydra-I cluster.I. Project description and preliminary results

Looking into the faintEst WIth MUSE (LEWIS) is an ESO large observing programme aimed at obtaining the first homogeneous integral-field spectroscopic survey of 30 extremely low-surface brightness (LSB) galaxies in the Hydra I cluster of galaxies, with MUSE at ESO-VLT. The majority of LSB galaxies in the sample (22 in total) are ultra-diffuse galaxies (UDGs). The distribution of systemic velocities Vsys ranges between 2317 km/s and 5198 km/s and is centred on the mean velocity of Hydra I (Vsys = 3683 $\pm$ 46 km/s). Considering the mean velocity and the velocity dispersion of the cluster, 17 out of 20 targets are confirmed cluster members. To assess the quality of the data and demonstrate the feasibility of the science goals, we report the preliminary results obtained for one of the sample galaxies, UDG11. For this target, we derived the stellar kinematics, including the 2-dimensional maps of line-of-sight velocity and velocity dispersion, constrained age and metallicity, and studied the globular cluster (GC) population hosted by the UDG. Results are compared with the available measurements for UDGs and dwarf galaxies in literature. By fitting the stacked spectrum inside one effective radius, we find that UDG11 has a velocity dispersion $\sigma = 20 \pm 8$ km/s, it is old ($10\pm1$ Gyr), metal-poor ([M/H]=-1.17$\pm$0.11 dex) and has a total dynamical mass-to-light ratio M$/L_V\sim 14$, comparable to those observed for classical dwarf galaxies. The spatially resolved stellar kinematics maps suggest that UDG11 does not show a significant velocity gradient along either major or minor photometric axes. We find two GCs kinematically associated with UDG11. The estimated total number of GCs in UDG11, corrected for the spectroscopic completeness limit, is $N_{GC}= 5.9^{+2.2}_ {-1.8}$, which corresponds to a GC specific frequency of $S_N = 8.4^{+3.2}_{-2.7}$.Comment: Accepted for publication in Astronomy and Astrophysic