G1/S restriction point controls phasic gene expression and differentiation during early mouse development.
Brian DeVeale, Leqian Liu, Ryan Boileau, Jennifer Swindlehurst-Chan, Bryan Marsh, Jake Freimer, Adam Abate and Robert Blelloch.
Nature Communications. June 2022. 13, 3696.
Pluripotent embryonic stem cells have a unique cell cycle structure with a suppressed G1/S restriction point and little differential expression across the cell cycle phases. Here, we evaluate the link between G1/S restriction point activation, phasic gene expression, and cellular differentiation. Expression analysis reveals a gain in phasic gene expression across lineages between embryonic days E7.5 and E9.5. Genetic manipulation of the G1/S restriction point regulators miR-302 and P27 respectively accelerates or delays the onset of phasic gene expression in mouse embryos. Loss of miR-302-mediated p21 or p27 suppression expedites embryonic stem cell differentiation, while a constitutive Cyclin E mutant blocks it. Together, these findings uncover a causal relationship between emergence of the G1/S restriction point with a gain in phasic gene expression and cellular differentiation.
tumorgenic non-coding mutations through altered cis-regulation.
Zhongshan Cheng, Michael Vermeulen, Micheal Rollins-Green, Tomas Babak,
STAR Protocols. Nov 2021. 2 (4), 100934.
Identification of non-coding mutations driving tumorigenesis requires alterna- tive approaches to coding mutations. Enriched associations between mutated regulatory elements and altered cis-regulation in tumors are a promising approach to stratify candidate non-coding driver mutations. Here we provide a bioinformatics pipeline to mine data from the Cancer Genomic Commons (GDC) for such associations. The pipeline integrates RNA and whole-genome sequencing with genotyping data to reveal putative non-coding driver mutations by cancer type.
mutations with driver hallmarks in major cancers.
Zhongshan Cheng, Michael Vermeulen, Micheal Rollins-Green, Brian DeVeale*,
iScience. Feb 2021. 24 (3), 102144.
*authors contributed equally
Despite the recent availability of complete genome sequences of tumors from thousands of patients, isolating disease-causing (driver) non-coding mutations from the plethora of somatic variants remains challenging, and only a handful of validated examples exist. By integrating whole-genome sequencing, genetic data, and allele-specific gene expression from TCGA, we identified 320 somatic non-coding mutations that affect gene expression in cis (FDR<0.25). These muta- tions cluster into 47 cis-regulatory elements that modulate expression of their subject genes through diverse molecular mechanisms. We further show that these mutations have hallmark features of non-coding drivers; namely, that they preferentially disrupt transcription factor binding motifs, are associated with a selective advantage, increased oncogene expression and decreased tumor suppressor expression.
microRNAs in mice.
Brian DeVeale, Jennifer Swindlehurst-Chan and Robert Blelloch. microRNAs in mice.
Nature Genetics Reviews.
Jan 2021. 22 (5), 307-323.
Hundreds of microRNAs (miRNAs) are expressed in distinct spatial and temporal patterns during embryonic and postnatal mouse development. The loss of all miRNAs through the deletion of critical miRNA biogenesis factors results in early lethality. The function of each miRNA stems from their cumulative negative regulation of multiple mRNA targets expressed in a particular cell type. During development, miRNAs often coordinate the timing and direction of cell fate transitions. In adults, miRNAs frequently contribute to organismal fitness through homeostatic roles in physiology. Here, we review how the recent dissection of miRNA-knockout phenotypes in mice as well as advances related to their targets, dosage, and interactions have collectively informed our understanding of the roles of miRNAs in mammalian development and adaptive responses.
Parental Bias has Benefits.
Brian DeVeale and Derek van der Kooy.
Neuron. Sept 2020. 107 (6), 994-996.
In this issue, Laukoter et al., 2020 report that parent-of-origin-dependent expression is homogeneous across distinct cortical cell types and within individual populations. Conversely, they observe preferential sensitivity of astrocytes to altered doses of imprinted loci.
Surfaceome profiling reveals novel regulators of neural stem cell function.
Brian DeVeale, Damaris Bausch-Fluck, Raewyn M. Seaberg, Susan Runciman, Vahe Akbarian, Phillip Karpowicz, Charles Yoon, Hannah Song, Rachel Leeder, Peter W. Zandstra, Bernd Wollscheid and Derek van der Kooy.
Stem Cells. 2014 Jan;32(1):258-68.
The composition of cell-surface proteins changes during lineage specification, altering cellular responses to their milieu. The changes that characterize maturation of early neural stem cells (NSCs) remain poorly understood. Here we use mass spectrometry-based cell surface capture technology to profile the cell surface of early NSCs and demonstrate functional requirements for several enriched molecules. Primitive NSCs arise from embryonic stem cells upon removal of Transforming growth factor-b signaling, while definitive NSCs arise from primitive NSCs upon Lif removal and FGF addition. In vivo aggregation assays revealed that N-cadherin upregulation is sufficient for the initial exclusion of definitive NSCs from pluripotent ectoderm, while c-kit signal- ing limits progeny of primitive NSCs. Furthermore, we implicate EphA4 in primitive NSC survival signaling and Erbb2 as being required for NSC proliferation. This work elucidates several key mediators of NSC function whose relevance is confirmed on forebrain-derived populations and identifies a host of other candidates that may regulate NSCs.
Oct4 is required ~E7.5 for
proliferation in the primitive streak.
Brian DeVeale, Irina Brokhman, Paria Mohseni, Tomas Babak, Charles Yoon, Anthony Lin, Kento Onishi, Alexey Tomilin, Larysa Pevny, Peter W. Zandstra, Andras Nagy and Derek van der Kooy.
PLoS Genetics. 2013 Nov;9(11):e1003957.
Oct4 is a widely recognized pluripotency factor as it maintains Embryonic Stem (ES) cells in a pluripotent state, and, in vivo, prevents the inner cell mass (ICM) in murine embryos from differentiating into trophectoderm. However, its function in somatic tissue after this developmental stage is not well characterized. Using a tamoxifen-inducible Cre recombinase and floxed alleles of Oct4, we investigated the effect of depleting Oct4 in mouse embryos between the pre-streak and headfold stages, ,E6.0–E8.0, when Oct4 is found in dynamic patterns throughout the embryonic compartment of the mouse egg cylinder. We found that depletion of Oct4 ,E7.5 resulted in a severe phenotype, comprised of craniorachischisis, random heart tube orientation, failed turning, defective somitogenesis and posterior truncation. Unlike in ES cells, depletion of the pluripotency factors Sox2 and Oct4 after E7.0 does not phenocopy, suggesting that ,E7.5 Oct4 is required within a network that is altered relative to the pluripotency network. Oct4 is not required in extraembryonic tissue for these processes, but is required to maintain cell viability in the embryo and normal proliferation within the primitive streak. Impaired expansion of the primitive streak occurs coincident with Oct4 depletion ,E7.5 and precedes deficient convergent extension which contributes to several aspects of the phenotype.
Critical evaluation of imprinted gene expression by RNA-
Seq: a new perspective.
Brian DeVeale, Derek van der Kooy and Tomas Babak.
PLoS Genetics. 2012;8(3):e1002600.
In contrast to existing estimates of approximately 200 murine imprinted genes, recent work based on transcriptome sequencing uncovered parent-of-origin allelic effects at more than 1,300 loci in the developing brain and two adult brain regions, including hundreds present in only males or females. Our independent replication of the embryonic brain stage, where the majority of novel imprinted genes were discovered and the majority of previously known imprinted genes confirmed, resulted in only 12.9% concordance among the novel imprinted loci. Further analysis and pyrosequencing-based validation revealed that the vast majority of the novel reported imprinted loci are false-positives explained by technical and biological variation of the experimental approach. We show that allele-specific expression (ASE) measured with RNA–Seq is not accurately modeled with statistical methods that assume random independent sampling and that systematic error must be accounted for to enable accurate identification of imprinted expression. Application of a robust approach that accounts for these effects revealed 50 candidate genes where allelic bias was predicted to be parent-of-origin–dependent. However, 11 independent validation attempts through a range of allelic expression biases confirmed only 6 of these novel cases. The results emphasize the importance of independent validation and suggest that the number of imprinted genes is much closer to the initial estimates.
Global survey of
genomic imprinting by transcriptome sequencing.
Babak T*, DeVeale B*, Armour C, Raymond C, Cleary MA, van der Kooy D, Johnson JM, Lim LP.
Current Biology. 2008 18: 1735-41.
*authors contributed equally
Genomic imprinting restricts gene expression to a paternal or maternal allele. To date, w90 imprinted transcripts have been identified in mouse, of which the majority were detected after intense interrogation of clusters of imprinted genes identified by phenotype-driven assays in mice with uniparen- tal disomies . Here we use selective priming and parallel sequencing to measure allelic bias in whole transcriptomes. By distinguishing parent-of-origin bias from strain-specific bias in embryos derived from a reciprocal cross of mice, we constructed a genome-wide map of imprinted transcription. This map was able to objectively locate over 80% of known imprinted loci and allowed the detection and confirmation of six novel imprinted genes. Even in the intensely studied embryonic day 9.5 developmental stage that we analyzed, more than half of all imprinted single-nucleotide polymor- phisms did not overlap previously discovered imprinted tran- scripts; a large fraction of these represent novel noncoding RNAs within known imprinted loci. For example, a previously unnoticed, maternally expressed antisense transcript was mapped within the Grb10 locus. This study demonstrates the feasibility of using transcriptome sequencing for map- ping of imprinted gene expression in physiologically normal animals. Such an approach will allow researchers to study imprinting without restricting themselves to individual loci or specific transcripts.
Immunity and Aging: the Enemy Within?
Brian DeVeale, Ted Brummel, Laurent Seroude.
Aging Cell, 2004 3:
Functional analyses of changes in the immune response indicate that aging is associated with a decline of adapt- ive immunity whereas innate immunity is ramped up. Gene expression studies also support age-dependent changes in immunity. Studies using a large panel of meth- odologies and multiple species show that some of the most dramatic transcriptional changes that occur during aging are associated with immunity. This observation leads to two fundamental questions: (1) Why is the immune response altered with age? (2) Is this a conse- quence of aging or does it contribute to it? The origin of these changes and the mechanistic relationship among them as well as with aging must be identified. In mam- mals, this task is complicated by the interdependence of the innate and adaptive immune systems. The value of invertebrates as model organisms to help answer these questions is presented. This includes a description of the immune response in invertebrate models and how it com- pares with vertebrates, focusing on conserved pathways. Finally, these questions are explored in light of recent reports and data from our laboratory. Experimental altera- tions of longevity indicate that the differential expres- sion of immunity-related genes during aging is linked to the rate of aging. Long-lived nematodes are more resist- ant to pathogens and blocking the expression of immune- related genes can prevent lifespan extension. These observations suggest that the immune response has a positive effect on longevity, possibly by increasing fit- ness. By contrast, it has been reported that activation of the immune system can reduce longevity upon starvation. We also observed that deregulation of the immune response has drastic effects on viability and longevity in Drosophila. These data suggest that the immune response results in a trade-off between beneficial and detrimental effects that might profoundly affect the aging process. Given this, immunity may be an ally early in life, but turns out to be an enemy as we age.