Noncommutative Biology: Sequential Regulation of Complex Networks and Connected Matter

The lin-12 locus specifies cell fates in caenorhabditis elegans

The regulation of gene expression is a key factor influencing cell fate decisions during differentiation. The molecular mechanisms underlying gene regulation are multi-layered, with regulatory inputs traversing both within and across different layers, thus providing a very difficult system to decipher. Although the gene expression patterns are cell-type specific, the regulatory inputs follow similar fundamental guidelines. One basic and easily interpretable transcriptional control layer is chromatin accessibility which is a consequence of combinatorial binding of transcription factors (TFs) and various chromatin modifications. Hematopoiesis is a well-characterized system for understanding transcriptional control mechanisms. The differentiation of hemogenic endothelium (HE) to hematopoietic progenitor cells (HP) forms a crucial step in hematopoiesis as it marks the exclusive commitment of bi-potent stem cells capable of following both hematopoietic and endothelial lineages, to hematopoietic lineages. In this work, we have performed a genome-wide analysis of chromatin accessibility dynamics in the differentiation of HE to HP and subsequently correlated them to gene expression dynamics for comprehending the basic regulatory themes. Chromatin accessibility was examined by global DNasel Hypersensitivity Sites (DHS) mapping during the differentiation, in addition to measuring gene expression by RNA-Seq for HE and HP. Accessible regions unique to HE and HP, and common peaks were identified. In general, there was a direct correlation between the DHS fold change and the expression fold change of the nearest genes for the corresponding peaks, with peaks near promoter regions showing a considerably higher degree of correlation than the rest of the peak populations. In further analyses, we applied a range of bioinformatics approaches such as peak overlap analysis, de novo motif discovery, and gene ontology analysis with the ultimate aim to identify the factors influencing lineage commitment. To this end, we have formulated a novel cluster-based approach for studying motif co-occurrence to obtain potential combinatorial TF binding regions, and by correlating those with the expression of the TFs we have systematically explored the relationship between the changes in chromatin accessibility and gene expression dynamics. We are presently developing mathematical models that integrate chromatin accessibility, chromatin modifications and transcription factors to predict gene expression patterns specific for a given cellular differentiation state.

Cell-type diversity is governed in part by differential gene expression programs mediated by transcription factor (TF) binding. However, there are few systematic studies of the genomic binding of different types of TFs across a wide range of human cell types, especially in relation to gene expression. In the ENCODE Project, we have identified the genomic binding locations across 11 different human cell types of CTCF, RNA Pol II (RNAPII), and MYC, three TFs with diverse roles. Our data and analysis revealed how these factors bind in relation to genomic features and shape gene expression and cell-type specificity. CTCF bound predominantly in intergenic regions while RNAPII and MYC preferentially bound to core promoter regions. CTCF sites were relatively invariant across diverse cell types, while MYC showed the greatest cell-type specificity. MYC and RNAPII co-localized at many of their binding sites and putative target genes. Cell-type specific binding sites, in particular for MYC and RNAPII, were associated with cell-type specific functions. Patterns of binding in relation to gene features were generally conserved across different cell types. RNAPII occupancy was higher over exons than adjacent introns, likely reflecting a link between transcriptional elongation and splicing. TF binding was positively correlated with the expression levels of their putative target genes, but combinatorial binding, in particular of MYC and RNAPII, was even more strongly associated with higher gene expression. These data illuminate how combinatorial binding of transcription factors in diverse cell types is associated with gene expression and cell-type specific biology.

Alternative Neural Crest Cell Fates Are Instructively Promoted by TGFβ Superfamily Members

Contribution of Bistability and Noise to Cell Fate Transitions Determined by Feedback Opening

A general interpolation method for constructing smooth molecular potential energy surfaces (PES’s) from ab initio data are proposed within the framework of the reproducing kernel Hilbert space and the inverse problem theory. The general expression for an a posteriori error bound of the constructed PES is derived. It is shown that the method yields globally smooth potential energy surfaces that are continuous and possess derivatives up to second order or higher. Moreover, the method is amenable to correct symmetry properties and asymptotic behavior of the molecular system. Finally, the method is generic and can be easily extended from low dimensional problems involving two and three atoms to high dimensional problems involving four or more atoms. Basic properties of the method are illustrated by the construction of a one-dimensional potential energy curve of the He–He van der Waals dimer using the exact quantum Monte Carlo calculations of Anderson et al. [J. Chem. Phys. 99, 345 (1993)], a two-dimensional potential energy surface of the HeCO van der Waals molecule using recent ab initio calculations by Tao et al. [J. Chem. Phys. 101, 8680 (1994)], and a three-dimensional potential energy surface of the H+3 molecular ion using highly accurate ab initio calculations of Röhse et al. [J. Chem. Phys. 101, 2231 (1994)]. In the first two cases the constructed potentials clearly exhibit the correct asymptotic forms, while in the last case the constructed potential energy surface is in excellent agreement with that constructed by Röhse et al. using a low order polynomial fitting procedure.

Bmgsb is involved in the determination of cell fate by affecting the cell cycle genes in the silk gland of Bombyx mori

Cell fate determination is usually described as the result of the stochastic dynamics of gene regulatory networks (GRNs) reaching one of multiple steady-states each of which corresponds to a specific decision. However, the fate of a cell is determined in finite time suggesting the importance of transient dynamics in cellular decision making. Here we consider cellular decision making as resulting from first passage processes of regulatory proteins and examine the effect of transient dynamics within the initial lysis-lysogeny switch of phage λ. Importantly, the fate of an infected cell depends, in part, on the number of coinfecting phages. Using a quantitative model of the phage λ GRN, we find that changes in the likelihood of lysis and lysogeny can be driven by changes in phage co-infection number regardless of whether or not there exists steady-state bistability within the GRN. Furthermore, two GRNs which yield qualitatively distinct steady state behaviors as a function of phage infection number can show similar transient responses, sufficient for alternative cell fate determination. We compare our model results to a recent experimental study of cell fate determination in single cell assays of multiply infected bacteria. Whereas the experimental study proposed a “quasi-independent” hypothesis for cell fate determination consistent with an observed data collapse, we demonstrate that observed cell fate results are compatible with an alternative form of data collapse consistent with a partial gene dosage compensation mechanism. We show that including partial gene dosage compensation at the mRNA level in our stochastic model of fate determination leads to the same data collapse observed in the single cell study. Our findings elucidate the importance of transient gene regulatory dynamics in fate determination, and present a novel alternative hypothesis to explain single-cell level heterogeneity within the phage λ lysis-lysogeny decision switch.

During development, neuronal identity is determined by a combination of numerous transcription factors. However, the mechanisms of synergistic action of these factors in transcriptional regulation and subsequent cell fate specification are largely unknown. In this study, we identified a novel gene, Corl1, encoding a nuclear protein with homology to the Ski oncoprotein. Corl1 was highly selectively expressed in the central nervous system (CNS). In the embryonic CNS, Corl1 was expressed in a certain subset of postmitotic neurons generated posterior to the midbrain-hindbrain border. In the developing spinal cord, Corl1 was selectively expressed in the dorsal horn interneurons where a homeodomain transcription factor, Lbx1, is required for proper specification. Corl1 was localized in a nuclear dot-like structure and interacted with general transcriptional corepressors. In addition, Corl1 showed transcriptional repression activity in the GAL4-fusion system, indicating its involvement in the regulation of transcriptional repression. Furthermore, Corl1 interacted with Lbx1 and cooperatively repressed transcription, suggesting that it acts as a transcriptional corepressor for Lbx1 in regulating cell fate determination in the dorsal spinal cord. Corl1 corepressor activity did not depend on Gro/TLE activity, and Gro/TLE also functioned as a corepressor for Lbx1. Thus, Lbx1 can select two independent partners, Corl1 and Gro/TLE, as corepressors. Identification of a novel transcriptional corepressor with neuronal subtype-restricted expression might provide insights into the mechanisms of cell fate determination in neurons.

Epidermal Growth Factor Receptor (EGFR) tyrosine kinase inhibitors (TKIs) are a first line treatment for patients with advanced non-small cell lung cancer (NSCLC) who have specific hyperactive mutations in EGFR. Though patients have seen improvements in survival after EGFR TKI treatment, resistance commonly develops as not all NSCLC cells die following treatment. Indeed, EGFR inhibitor treatment induces both cell-cycle arrest (allowing for cell survival) and death in cancer cells. The FOXO family of transcription factors have been implicated in both death and arrest responses following EGFR inhibition. This is consistent with FOXOs function in regulating both arrest and apoptotic genes programs. However, it is unknown how the FOXO network ‘decides' between these two oppositional programs to enact following treatment with EGFR inhibitors. Protein dynamics, meaning how the levels or location of a protein changes over time, is a mechanism cells use to encode information about cellular fates. The p53 transcription system is an established example of a dynamically regulated system–it responds to multiple inputs and is capable of enacting multiple outputs (gamma radiation causes oscillations of p53 levels that result in cellular arrest while UV causes a sustained increase of p53 levels that result in cellular death). FOXO's network is similar to the p53 system (multiple input, multiple output), making it a good candidate for dynamic regulation. We have tagged Foxo3a with a fluorescent marker (Venus) at its endogenous locus in PC9 NSCLC cells using CRISPR/Cas9 to determine whether FOXO dynamics dictate cell fate following EGFR TKI treatment. Using live cell microscopy on this line, we have acquired FOXO nuclear cytoplasmic shuttling dynamics as well as cell fate (death/arrest) in single cells. Using time-series analyses we are trying to determine if there are dynamic patterns associated with each fate following EGFR inhibition. Through our studies, we hope to gain a better understanding of this non-genetic mechanism cells use to determine their fate. Our long-term goal is to identify dynamic patterns that push cancer cells to more terminal fates following treatment. Citation Format: Julie M. Huynh, Kayenat S. Aryeh, Lisa Shanks, Andrew L. Paek. Role of FOXO transcription factor dynamics in determining cell fates following EGFR inhibition [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2471.

Dynamic Filopodia Transmit Intermittent Delta-Notch Signaling to Drive Pattern Refinement during Lateral Inhibition

The Protean Nature of Cells in the B Lymphocyte Lineage

Primitive endoderm (PE) and epiblast (EPI) are two lineages derived from the inner cell mass (ICM) of the E3.5 blastocyst. Recent studies showed that EPI and PE progenitors expressing the lineage-specific transcriptional factors Nanog and Gata6, respectively, arise progressively as the ICM develops. Subsequent sorting of the two progenitors during blastocyst maturation results in the ormation of morphologically distinct EPI and PE layers at E4.5. It is, however, unknown how the initial differences between the two populations become established in the E3.5 blastocyst. Because the ICM cells are derived from two distinct rounds of polarized cell divisions during cleavage, a possible role for cell lineage history in promoting EPI versus PE fate has been proposed. We followed cell lineage from the eight-cell stage by live cell tracing and could find no clear linkage between developmental history of individual ICM cells and later cell fate. However, modulating FGF signaling levels by inhibition of the receptor/MAP kinase pathway or by addition of exogenous FGF shifted the fate of ICM cells to become either EPI or PE, respectively. Nanog- or Gata6-expressing progenitors could still be shifted towards the alternative fate by modulating FGF signaling during blastocyst maturation, suggesting that the ICM progenitors are not fully committed to their final fate at the time that initial segregation of gene expression occurs. In conclusion, we propose a model in which stochastic and progressive specification of EPI and PE lineages occurs during maturation of the blastocyst in an FGF/MAP kinase signal-dependent manner.

Suppression of Ptf1a Activity Induces Acinar-to-Endocrine Conversion

Lung development relies on diverse cell intrinsic and extrinsic mechanisms to ensure proper cellular differentiation and compartmentalization. In addition, it requires precise integration of multiple signaling pathways to temporally regulate morphogenesis and appropriate cell specification. To accomplish this, organogenesis relies on epigenetic and transcriptional regulators to promote cell fate and inhibit alternative cell fates. Using genetic mouse and human embryonic stem cell (hESC) differentiation models, tissue explants, and single-cell transcriptomic analysis, we demonstrated that Bromodomain Containing Protein 4 (BRD4) is required for mammalian lung morphogenesis and cell fate. Endodermal deletion of BRD4 impaired epithelial-mesenchymal crosstalk, leading to disrupted proximal-distal patterning and branching morphogenesis. Moreover, temporal deletion of BRD4 revealed developmental stage–specific defects in airway and alveolar epithelial cell specification with a predominant role in proximal airway cell fate. Similarly, BRD4 promoted lung endodermal cell differentiation into airway lineages in a hESC-derived lung organoid model. Together, these data demonstrate that BRD4 orchestrates early lung morphogenesis and separately regulates cell specification, indicating a multifunctional and evolutionarily conserved role for BRD4 in mammalian lung development.