Cellular Developmental Programs Research Articles

Human prefrontal cortex gene regulatory dynamics from gestation to adulthood at single-cell resolution

Human brain development is underpinned by cellular and molecular reconfigurations continuing into the third decade of life. To reveal cell dynamics orchestrating neural maturation, we profiled human prefrontal cortex gene expression and chromatin accessibility at single-cell resolution from gestation to adulthood. Integrative analyses define the dynamic trajectories of each cell type, revealing major gene expression reconfiguration at the prenatal-to-postnatal transition in all cell types followed by continuous reconfiguration into adulthood and identifying regulatory networks guiding cellular developmental programs, states, and functions. We uncover links between expression dynamics and developmental milestones, characterize the diverse timing of when cells acquire adult-like states, and identify molecular convergence from distinct developmental origins. We further reveal cellular dynamics and their regulators implicated in neurological disorders. Finally, using this reference, we benchmark cell identities and maturation states in organoid models. Together, this captures the dynamic regulatory landscape of human cortical development.

Pulsed electromagnetic fields synergize with graphene to enhance dental pulp stem cell-derived neurogenesis by selectively targeting TRPC1 channels

Conventional root canal treatment replaces the infected pulp with defined materials. Alternative cell-based tissue engineering strategies aim to regenerate a fully functional pulp within the root canal. Despite recent advances in this area, however, the regeneration of an innervated pulp remains a major challenge in the field. Both graphene (2DG) and pulsed electromagnetic fields (PEMFs) independently have been shown to promote diverse cellular developmental programs. The present study showed that 2DG promoted the neurogenic induction of human dental pulp stem cells (hDPSCs) by upregulating and accelerating the expression of mature neuronal markers. Notably, 2DG induced the highest expression of transient receptor potential canonical cation channel type 1 (TRPC1) during early neurogenesis. As brief PEMF exposure promotes in vitro differentiation by activating a TRPC1-mitochondrial axis, an opportunity to combine 2DG with developmentally targeted PEMF exposure for synergistic effects was realizable. Neurogenic gene expression, neurotransmitter release, and reactive oxygen species (ROS) production were greatly enhanced by a brief (10 min) and low amplitude (2 mT) PEMF exposure timed to coincide with the highest TRPC1 expression from hDPSCs on 2DG. In contrast, hDPSCs on glass were less responsive to PEMF exposure. The capacity of PEMFs to promote neurogenesis was precluded by the administration of penicillin/streptomycin, mirroring previous studies demonstrating that aminoglycoside antibiotics block TRPC1-mediated calcium entry and verifying the contribution of TRPC1 in this form of magnetoreception. Hence, graphene created a more conducive environment for subsequent PEMF-stimulated neurogenic induction of hDPSCs through their mutual capacity to activate TRPC1with subsequent ROS production.

Matrix Production and Sporulation in Bacillus subtilis Biofilms Localize to Propagating Wave Fronts

Bacterial biofilms are surface-attached microbial communities encased in self-produced extracellular polymeric substances. Here we demonstrate that during the development of Bacillus subtilis biofilms, matrix production is localized to an annular front propagating at the periphery and sporulation to a second front at a fixed distance at the interior. We show that within these fronts, cells switch off matrix production and transition to sporulation after a set time delay of ∼100 min. Correlation analyses of fluctuations in fluorescence reporter activity reveal that the fronts emerge from a pair of gene-expression waves of matrix production and sporulation. The localized expression waves travel across cells that are immobilized in the biofilm matrix in contrast to active cell migration or horizontal colony spreading. Our results suggest that front propagation arises via a local developmental program occurring at the level of individual bacterial cells, likely driven by nutrient depletion and metabolic by-product accumulation. A single-length scale and timescale couples the spatiotemporal propagation of both fronts throughout development. As a result, gene expression patterns within the advancing fronts collapse to self-similar expression profiles. Our findings highlight the key role of the localized cellular developmental program associated with the propagating front in describing biofilm growth.

Circuitry Rewiring Directly Couples Competence to Predation in the Gut Dweller Streptococcus salivarius

Small distortions in transcriptional networks might lead to drastic phenotypical changes, especially in cellular developmental programs such as competence for natural transformation. Here, we report a pervasive circuitry rewiring for competence and predation interplay in commensal streptococci. Canonically, in streptococci paradigms such as Streptococcus pneumoniae and Streptococcus mutans, the pheromone-based two-component system BlpRH is a central node that orchestrates the production of antimicrobial compounds (bacteriocins) and incorporates signal from the competence activation cascade. However, the human commensal Streptococcus salivarius does not contain a functional BlpRH pair, while the competence signaling system ComRS directly couples bacteriocin production and competence commitment. This network shortcut might underlie an optimal adaptation against microbial competitors and explain the high prevalence of S.salivarius in the human digestive tract. Moreover, the broad spectrum of bacteriocin activity against pathogenic bacteria showcases the commensal and genetically tractable S.salivarius species as a user-friendly model for competence and bacterial predation.

Polycomb and trithorax opposition in development and disease.

Early discoveries in chromatin biology and epigenetics heralded new insights into organismal development. From these studies, two mediators of cellular differentiation were discovered: the Polycomb group (PcG) of transcriptional repressors, and the trithorax group (trxG) of transcriptional activators. These protein families, while opposed in function, work together to coordinate the appropriate cellular developmental programs that allow for both embryonic stem cell self-renewal and differentiation. Recently, both the PcG and trxG chromatin modulators have been observed to be deregulated in a wide spectrum diseases including developmental disorders and cancer. To understand the impact of these findings we outline the past, present, and future. WIREs Dev Biol 2016, 5:659-688. doi: 10.1002/wdev.244 For further resources related to this article, please visit the WIREs website.

Activation of the Hedgehog Signaling Pathway in the Developing Lens Stimulates EctopicFoxE3Expression and Disruption in Fiber Cell Differentiation

The signaling pathways and transcriptional effectors responsible for directing mammalian lens development provide key regulatory molecules that can inform our understanding of human eye defects. The hedgehog genes encode extracellular signaling proteins responsible for patterning and tissue formation during embryogenesis. Signal transduction of this pathway is mediated through activation of the transmembrane proteins smoothened and patched, stimulating downstream signaling resulting in the activation or repression of hedgehog target genes. Hedgehog signaling is implicated in eye development, and defects in hedgehog signaling components have been shown to result in defects of the retina, iris, and lens. We assessed the consequences of constitutive hedgehog signaling in the developing mouse lens using Cre-LoxP technology to express the conditional M2 smoothened allele in the embryonic head and lens ectoderm. Although initial lens development appeared normal, morphological defects were apparent by E12.5 and became more significant at later stages of embryogenesis. Altered lens morphology correlated with ectopic expression of FoxE3, which encodes a critical gene required for human and mouse lens development. Later, inappropriate expression of the epithelial marker Pax6, and as well as fiber cell markers c-maf and Prox1 also occurred, indicating a failure of appropriate lens fiber cell differentiation accompanied by altered lens cell proliferation and cell death. Our findings demonstrate that the ectopic activation of downstream effectors of the hedgehog signaling pathway in the mouse lens disrupts normal fiber cell differentiation by a mechanism consistent with a sustained epithelial cellular developmental program driven by FoxE3.

DNMT3A and stem cell function: new insights into old pathways

DNA methylation represents an important epigenetic regulator of gene expression which impacts cellular developmental programs through alterations in gene expression. DNA methylation is established by highly related de novo DNA methyltransferases Dnmt3a and Dnmt3b, and maintained by Dnmt1.[1][1]

Regulation of Central Metabolism Genes of Mycobacterium tuberculosis by Parallel Feed-Forward Loops Controlled by Sigma Factor E (σ E )

Cells respond to external stimuli through networks of regulatory interactions. The human pathogen Mycobacterium tuberculosis responds to stress encountered during infection by arresting multiplication and implementing critical metabolic changes that lead to or sustain the nonreplicative state. Much of this differentiation program is recapitulated when M. tuberculosis cultures are subjected to gradual oxygen depletion in vitro. Here we report that hypoxic induction of critical central metabolism genes in the glyoxylate shunt (icl1) and in the methylcitrate cycle (gltA1) involves both global and local regulators. The global regulators are accessory sigma factors σ(B) for icl1 and σ(E) for gltA1. The local regulators are the products of two paralogous genes mapping at positions adjacent to the corresponding effector gene or operon. We call these genes lrpI and lrpG (for local regulatory protein of icl1 and gltA1). We also found that (i) each sigma factor controls the corresponding local regulator, (ii) both global and local regulators are required for effector gene induction, and (iii) the occurrence of sigma factor control of effector gene induction is independent of its control over the corresponding local regulator. Together, these data indicate that induction of icl1 and gltA1 utilizes parallel feed-forward loops with an AND input function. Both feed-forward loops are affected by σ(E), since this sigma factor is part of the gltA1 loop and controls sigB in the icl1 loop. Feed-forward loops may critically contribute to the cellular developmental program associated with M. tuberculosis dormancy.

Engrafting fetal liver cells into multiple tissues of healthy adult mice without the use of immunosuppressants

We have shown the fetal liver cell engraftments into multiple tissues of adult healthy mice, achieved without suppressing the animals’ immune systems. Fetal cells from the livers of male C57Bl/6J Black lineage mice at day 13 to 15 of gestation were injected intravenously into female adult CC57W/MY White mice. The grafting was evaluated by Y-chromosome-specific PCR, cytometric analysis of fluorescently stained donor cells, and histological analysis. All the methods consistently showed the presence of multiple engraftments randomly distributed through the various organs of the recipients. After 60 days, the grafts still constituted 0.1 to 2.75% of the tissues. The grafted cells did not change their appearance in any of the organs except the brain, where they became enlarged. Inflammatory reactions were not detected in any of the histological preparations. The frequency of engraftments was higher in the liver, indicating that similarity between the donor and recipient cells facilitates engraftment. The high inherent plasticity of fetal liver cells underlies their ability to integrate into healthy recipient organs, which can be governed by environmental conditions and connections with neighboring cells rather than by the initial cellular developmental programs. The fact that fetal liver cells can be grafted into multiple tissues of healthy animals indicates that they can be used to replace the natural loss of cells in adult organisms.

Expression of MLL Fusion Genes in Human Hematopoietic Cells Is Sufficient To Induce Both Acute Lymphoid and Acute Myeloid Leukemia In Vivo.

Malignant transformation involves the acquisition of a series of genetic and epigenetic changes that subvert the normal cellular developmental program and result in the generation of a neoplastic clone with deregulated growth properties. However, in some instances, acquisition of a single oncogenic hit may be sufficient to induce a clinical disease. Studies on identical twins that developed acute leukemias during their childhood have clearly demonstrated that all oncogenes are not equal with some exhibiting greater potency than others. MLL translocations, the most frequent cause of infant leukemias, occur in utero and can be identified on Guthrie cards. Identical twins have a concordance rate near 100% with a short latency, a near synchronous diagnosis and a very poor prognosis, suggesting that MLL leukemias are a single hit disease.In order to test the effects and potency of MLL fusion genes in human hematopoietic cells, we conducted a series of in vivo experiments using retroviruses encoding either MLL-ENL or MLL-AF9. Lineage-depleted cord blood cells were transduced with these viruses then injected into sub-lethally irradiated NOD/SCID mice. 15 out of 16 MLL-ENL mice developed an aggressive human pro-B acute lymphoblastic leukemia (ALL) characterized by the extensive accumulation of immature B cells in the bone marrow and organs in less than 18 weeks. 5 out of 12 MLL-AF9 mice developed the same B-precursor ALL while 2 others developed myelomonocytic leukemia. 5 of the 6 mice injected with MLL-transduced cells did not show evidence of viral integration by PCR. The overall penetrance of leukemia in primary mice with evidence of viral integration was greater than 95%. Transplantation of leukemic cells from primary mice to secondary recipients was able to recapitulate the disease with the same phenotype in a shorter period of time with a penetrance of 100%. Karyotypic analysis of leukemic blasts from primary and secondary mice was normal.Clonal analysis of retroviral integration showed that leukemias generated in primary mice were either monoclonal or oligoclonal (two dominating clones) while the majority of secondary mice had monoclonal disease. At the level of the immunoglobulin (IgH) rearrangements, the B-precursor leukemias were always polyclonal in primary mice, with the presence of the germline rearrangement in addition to other rearrangements. New clones appeared in secondary recipients, suggesting constant rearrangement of the IgH locus. These data also support that a more primitive cell type that had not yet rearranged its immunoglobulin genes was the target of the transforming event, ruling out the possibility that a mature B cell is the target of the transformation.To our knowledge, these results provide the first examples of in vivo models where human hematopietic stem/progenitor cells are transformed into leukemia, recapitulating the features of the human disease. Also, this represents the first reliable in vivo model of B-precursor ALL using MLL fusion genes and highlights species-specific differences in the mechanisms of neoplastic transformation of human and murine cells. The very high penetrance, short latency, normal karyotype and multiple clones that contribute to the disease collectively demonstrate that expression of MLL fusions is sufficient to induce human leukemias in vivo and support the notion that the MLL leukemias are the result of a single genetic hit.