Structure of a bacterial ribonucleoprotein complex central to the control of cell envelope biogenesis

The biogenesis of the essential precursor of the bacterial cell envelope, glucosamine-6-phosphate (GlcN6P), is controlled through intricate post-transcription networks mediated by GlmZ, a small regulatory RNA (sRNA). GlmZ stimulates translation of the mRNA encoding GlcN6P synthetase in Escherichia coli, but when bound by the protein RapZ, it becomes inactivated through cleavage by the endoribonuclease RNase E. Here we report the cryoEM structure of the RapZ:GlmZ complex, revealing a complementary match of the protein tetrameric quaternary structure to an imperfect structural repeat in the RNA. The RNA is contacted mostly through a highly conserved domain of RapZ that shares deep evolutionary relationship with phosphofructokinase and suggests links between metabolism and riboregulation. We also present the structure of a pre-cleavage encounter intermediate formed between the binary RapZ:GlmZ complex and RNase E that reveals how GlmZ is presented and recognised for cleavage. The structures suggest how other encounter complexes might guide recognition and action of endoribonucleases on target transcripts, and how structured substrates in polycistronic precursors are recognised for processing.

TAF4b transcription networks regulating early oocyte differentiation

Establishment of a healthy ovarian reserve is contingent upon numerous regulatory pathways during embryogenesis. Previously, mice lacking TBP-associated factor 4b (Taf4b) were shown to exhibit a diminished ovarian reserve. However, potential oocyte-intrinsic functions of TAF4b have not been examined. Here we use a combination of gene expression profiling and chromatin mapping to characterize the TAF4b gene regulatory network in mouse oocytes. We find that Taf4b-deficient oocytes display inappropriate expression of meiotic, chromatin, and X-linked genes, and unexpectedly we found a connection with Turner Syndrome pathways. Using Cleavage Under Targets and Release Using Nuclease (CUT&RUN), we observed TAF4b enrichment at genes involved in meiosis and DNA repair, some of which are differentially expressed in Taf4b-deficient oocytes. Interestingly, TAF4b target genes were enriched for Sp/KLF family motifs rather than TATA-box, suggesting an alternate mode of promoter interaction. Together, our data connects several gene regulatory nodes that contribute to the ovarian reserve.

Transcriptional profile in rat muscle: down-regulation networks in acute strenuous exercise

Background Physical exercise is a health promotion factor regulating gene expression and causing changes in phenotype, varying according to exercise type and intensity. Acute strenuous exercise in sedentary individuals appears to induce different transcriptional networks in response to stress caused by exercise. The objective of this research was to investigate the transcriptional profile of strenuous experimental exercise. Methodology RNA-Seq was performed with Rattus norvegicus soleus muscle, submitted to strenuous physical exercise on a treadmill with an initial velocity of 0.5 km/h and increments of 0.2 km/h at every 3 min until animal exhaustion. Twenty four hours post-physical exercise, RNA-seq protocols were performed with coverage of 30 million reads per sample, 100 pb read length, paired-end, with a list of counts totaling 12816 genes. Results Eighty differentially expressed genes (61 down-regulated and 19 up-regulated) were obtained. Reactome and KEGG database searches revealed the most significant pathways, for down-regulated gene set, were: PI3K-Akt signaling pathway, RAF-MAP kinase, P2Y receptors and Signaling by Erbb2. Results suggest PI3K-AKT pathway inactivation by Hbegf, Fgf1 and Fgr3 receptor regulation, leading to inhibition of cell proliferation and increased apoptosis. Cell signaling transcription networks were found in transcriptome. Results suggest some metabolic pathways which indicate the conditioning situation of strenuous exercise induced genes encoding apoptotic and autophagy factors, indicating cellular stress. Conclusion Down-regulated networks showed cell transduction and signaling pathways, with possible inhibition of cellular proliferation and cell degeneration. These findings reveal transitory and dynamic process in cell signaling transcription networks in skeletal muscle after acute strenuous exercise.

Suboptimal Intermediates Underlie Evolution of the Bicoid Homeodomain

Changes in regulatory networks generate materials for evolution to create phenotypic diversity. For transcription networks, multiple studies have shown that alterations in binding sites of cis-regulatory elements correlate well with the gain or loss of specific features of the body plan. Less is known about alterations in the amino acid sequences of the transcription factors (TFs) that bind these elements. Here we study the evolution of Bicoid (Bcd), a homeodomain (HD) protein that is critical for anterior embryo patterning in Drosophila. The ancestor of Bcd (AncBcd) emerged after a duplication of a Zerknullt (Zen)-like ancestral protein (AncZB) in a suborder of flies. AncBcd diverged from AncZB, gaining novel transcriptional and translational activities. We focus on the evolution of the HD of AncBcd, which binds to DNA and RNA, and is comprised of four subdomains: an N-terminal arm (NT) and three helices; H1, H2, and Recognition Helix (RH). Using chimeras of subdomains and gene rescue assays in Drosophila, we show that robust patterning activity of the Bcd HD (high frequency rescue to adulthood) is achieved only when amino acid substitutions in three separate subdomains (NT, H1, and RH) are combined. Other combinations of subdomains also yield full rescue, but with lower penetrance, suggesting alternative suboptimal activities. Our results suggest a multi-step pathway for the evolution of the Bcd HD that involved intermediate HD sequences with suboptimal activities, which constrained and enabled further evolutionary changes. They also demonstrate critical epistatic forces that contribute to the robust function of a DNA-binding domain.

Arabidopsis bZIP11 is a susceptibility factor during Pseudomonas syringae infection

The induction of plant nutrient secretion systems is critical for successful pathogen infection. Some bacterial pathogens, e.g. Xanthomonas species, use TAL (transcription activator-like) effectors to induce transcription of SWEET sucrose efflux transporters. Pseudomonas syringae pathovar (pv.) tomato strain DC3000 lacks TAL effectors, yet is able to induce multiple SWEETs in Arabidopsis thaliana by unknown mechanisms. Since bacteria require other nutrients besides sugars for efficient reproduction, we hypothesized that Pseudomonas may depend on host transcription factors involved in secretory programs to increase access to essential nutrients. Bioinformatic analyses identified the Arabidopsis basic-leucine zipper transcription factor bZIP11 as a potential regulator of nutrient transporters, including SWEETs and UmamiT amino acid transporters. Inducible downregulation of bZIP11 expression in Arabidopsis resulted in reduced growth of P. syringae pv. tomato strain DC3000, whereas inducible overexpression of bZIP11 resulted in increased bacterial growth, supporting the hypothesis that bZIP11 regulated transcription programs are essential for maximal pathogen titer in leaves. Our data are consistent with a model in which a pathogen alters host transcription factor expression upstream of secretory transcription networks to promote nutrient efflux from host cells.

Synergism and Antagonism of Two Distinct, but Confused, Nrf1 Factors in Integral Regulation of the Nuclear-to-Mitochondrial Respiratory and Antioxidant Transcription Networks

There is hitherto no literature available for explaining two distinct, but confused, Nrf1 transcription factors, because they shared the same abbreviations from nuclear factor erythroid 2-related factor 1 (also called Nfe2l1) and nuclear respiratory factor (originally designated α-Pal). Thus, we have here identified that Nfe2l1Nrf1 and α-PalNRF1 exert synergistic and antagonistic roles in integrative regulation of the nuclear-to-mitochondrial respiratory and antioxidant transcription profiles. In mouse embryonic fibroblasts (MEFs), knockout of Nfe2l1-/- leads to substantial decreases in expression levels of α-PalNRF1 and Nfe2l2, together with TFAM (mitochondrial transcription factor A) and other target genes. Similar inhibitory results were determined in Nfe2l2-/- MEFs but with an exception that both GSTa1 and Aldh1a1 were distinguishably upregulated in Nfe2l1-/- MEFs. Such synergistic contributions of Nfe2l1 and Nfe2l2 to the positive regulation of α-PalNRF1 and TFAM were validated in Keap1-/- MEFs. However, human α-PalNRF1 expression was unaltered by hNfe2l1α-/-, hNfe2l2-/-ΔTA, or even hNfe2l1α-/-+siNrf2, albeit TFAM was activated by Nfe2l1 but inhibited by Nfe2l2; such an antagonism occurred in HepG2 cells. Conversely, almost all of mouse Nfe2l1, Nfe2l2, and cotarget genes were downexpressed in α-PalNRF1+/- MEFs. On the contrary, upregulation of human Nfe2l1, Nfe2l2, and relevant reporter genes took place after silencing of α-PalNRF1, but their downregulation occurred upon ectopic expression of α-PalNRF1. Furtherly, Pitx2 (pituitary homeobox 2) was also identified as a direct upstream regulator of Nfe2l1 and TFAM, besides α-PalNRF1. Overall, these across-talks amongst Nfe2l1, Nfe2l2, and α-PalNRF1, along with Pitx2, are integrated from the endoplasmic reticulum towards the nuclear-to-mitochondrial communication for targeting TFAM, in order to finely tune the robust balance of distinct cellular oxidative respiratory and antioxidant gene transcription networks, albeit they differ between the mouse and the human. In addition, it is of crucial importance to note that, in view of such mutual interregulation of these transcription factors, much cautions should be severely taken for us to interpret those relevant experimental results obtained from knockout of Nfe2l1, Nfe2l2, α-Pal or Pitx2, or their gain-of-functional mutants.

Comparative Analysis of the Lignification Process of Two Bamboo Shoots Stored at Room Temperature

Two types of bamboo shoots, high bamboo (Phyllostachys prominens) shoots (HBSes) and moso bamboo (Phyllostachys edulis) shoots (MBSes), underwent a fast post-harvest lignification process under room temperature storage. To explore the mechanism of lignification in two types of bamboo shoots after post-harvest during room temperature storage, the measurement of cell wall polymers (lignin and cellulose) and enzyme activities of phenylalanine ammonialyase (PAL) and peroxidase (POD), and relative expression of related transcription networks factors (TFs) were performed. The results suggested that the lignification process in HBSes is faster than that in MBSes because of incremental increase in lignin and cellulose contents within 6 days and the shorter shelf-life. Additionally, compared with the expression pattern of lignification-related TFs and correlation analysis of lignin and cellulose contents, MYB20, MYB43, MYB85 could function positively in the lignification process of two types of bamboo shoots. A negative regulator, KNAT7, could negatively regulate the lignin biosynthesis in two types of bamboo shoots. In addition, MYB63 could function positively in HBSes, and NST1 could function negatively in MBSes. Notably, MYB42 may function differently in the two types of bamboo shoots, that is, a positive regulator in HBSes, but a negative regulator in MBSes. Transcription networks provide a comprehensive analysis to explore the mechanism of lignification in two types of bamboo shoots after post-harvest during room temperature storage. These results suggest that the lignification of bamboo shoots was mainly due to the increased activity of POD, higher expression levels of MYB20, MYB43, MYB63, and MYB85 genes, and lower expression levels of KNAT7 and NST1 genes, and the lignification process of HBSes and MBSes had significant differences.

Suboptimal intermediates underlie evolution of the Bicoid homeodomain

Changes in regulatory networks generate materials for evolution to create phenotypic diversity. For transcription networks, multiple studies have shown that alterations in binding sites of cis-regulatory elements correlate well with the gain or loss of specific features of the body plan. Less is known about alterations in the amino acid sequences of the transcription factors (TFs) that bind these elements. Here we study the evolution of Bicoid (Bcd), a homeodomain (HD) protein that is critical for anterior embryo patterning in Drosophila. The ancestor of Bcd (AncBcd) emerged after a duplication of a Zerknullt (Zen)-like ancestral protein (AncZB) in a suborder of flies. AncBcd diverged from AncZB, gaining novel transcriptional and translational activities. We focus on the evolution of the HD of AncBcd, which binds DNA and RNA, and is comprised of four subdomains: an N-terminal arm (NT) and three helices; H1, H2, and Recognition Helix (RH). Using chimeras of subdomains and gene rescue assays in Drosophila, we show that robust patterning activity of the Bcd HD (high frequency rescue to adulthood) is achieved only when amino acid substitutions in three separate subdomains (NT, H1, and RH) are combined. Other combinations of subdomains also yield full rescue, but with lower penetrance, suggesting alternative suboptimal activities. Our results suggest a multi-step pathway for the evolution of the Bcd HD that involved intermediate HD sequences with suboptimal activities, which constrained and enabled further evolutionary changes. They also demonstrate critical epistatic forces that contribute to the robust function of a DNA-binding domain.

Primate-restricted KRAB zinc finger proteins and target retrotransposons control gene expression in human neurons

In the first days of embryogenesis, transposable element–embedded regulatory sequences (TEeRS) are silenced by Kruppel-associated box (KRAB) zinc finger proteins (KZFPs). Many TEeRS are subsequently co-opted in transcription networks, but how KZFPs influence this process is largely unknown. We identify ZNF417 and ZNF587 as primate-specific KZFPs repressing HERVK (human endogenous retrovirus K) and SVA (SINE-VNTR-Alu) integrants in human embryonic stem cells (ESCs). Expressed in specific regions of the human developing and adult brain, ZNF417/587 keep controlling TEeRS in ESC-derived neurons and brain organoids, secondarily influencing the differentiation and neurotransmission profile of neurons and preventing the induction of neurotoxic retroviral proteins and an interferon-like response. Thus, evolutionarily recent KZFPs and their TE targets partner up to influence human neuronal differentiation and physiology.