Pharyngeal mesoderm regulatory network controls cardiac and head muscle morphogenesis

Harel, Itamar et al.The search for developmental mechanisms driving vertebrate organogenesis has paved the way toward a deeper understanding of birth defects. During embryogenesis, parts of the heart and craniofacial muscles arise from pharyngeal mesoderm (PM) progenitors. Here, we reveal a hierarchical regulatory network of a set of transcription factors expressed in the PM that initiates heart and craniofacial organogenesis. Genetic perturbation of this network in mice resulted in heart and craniofacial muscle defects, revealing robust cross-regulation between its members. We identified Lhx2 as a previously undescribed player during cardiac and pharyngeal muscle development. Lhx2 and Tcf21 genetically interact with Tbx1, the major determinant in the etiology of DiGeorge/velo-cardio-facial/22q11.2 deletion syndrome. Furthermore, knockout of these genes in the mouse recapitulates specific cardiac features of this syndrome. We suggest that PM-derived cardiogenesis and myogenesis are network properties rather than properties specific to individual PM members. These findings shed new light on the developmental underpinnings of congenital defects.This work was supported by grants to E.T. from the European Research Council; Israel Science Foundation; United States-Israel Binational Science Foundation; German Israeli Foundation; Association Française Contre les Myopathies; Kirk Center for Childhood Cancer and Immunological Disorders; Jeanne and Joseph Nissim Foundation for Life Sciences Research; and a donation from the Jack Gitlitz Estate. CK was supported by NIH-NIAMS grant AR054406. J.W.C was supported by a Studentship from The Institute Of Cancer Research, London. J.J.C. was partly supported by a Ministry of Science and Innovation (MICINN) grant [BFU2011-22928].Peer reviewe

Dial M(RF) for myogenesis

The transcriptional regulatory network that controls the determination and differentiation of skeletal muscle cells in the embryo has at its core the four myogenic regulatory factors (MRFs) Myf5, MyoD, Mrf4 and MyoG. These basic helix–loop–helix transcription factors act by binding, as obligate heterodimers with the ubiquitously expressed E proteins, to the E-box sequence CANNTG. While all skeletal muscle cells have the same underlying function their progenitors arise at many sites in the embryo and it has become apparent that the upstream activators of the cascade differ in these various populations so that it can be switched on by a variety of inductive signals, some of which act by initiating transcription, some by maintaining it. The application of genome-wide approaches has provided important new information as to how the MRFs function to activate the terminal differentiation programme and some of these data provide significant mechanistic insights into questions which have exercised the field for many years. We also consider the emerging roles played by micro-RNAs in the regulation of both upstream activators and terminal differentiation genes.Peer reviewe

A BAC transgenic analysis of the Mrf4/Myf5 locus reveals interdigitated elements that control activation and maintenance of gene expression during muscle development

The muscle-specific transcription factors Myf5 and Mrf4 are two of the four myogenic regulatory factors involved in the transcriptional cascade responsible for skeletal myogenesis in the vertebrate embryo. Myf5 is the first of these four genes to be expressed in the mouse. We have previously described discrete enhancers that drive Myf5 expression in epaxial and hypaxial somites, branchial arches and central nervous system, and argued that additional elements are required for proper expression (Summerbell, D., Ashby, P.R., Coutelle, O., Cox, D., Yee, S.P. and Rigby, P.W.J. (2000) Development 127, 3745-3757). We have now investigated the transcriptional regulation of both Myf5 and Mrf4 using bacterial artificial chromosome transgenesis. We show that a clone containing Myf5 and 140 kb of upstream sequences is sufficient to recapitulate the known expression patterns of both genes. Our results confirm and reinforce the conclusion of our earlier studies, that Myf5 expression is regulated differently in each of a considerable number of populations of muscle progenitors, and they begin to illuminate the evolutionary origins of this complex regulation. We further show that separate elements are involved in the activation and maintenance of expression in the various precursor populations, reflecting the diversity of the signals that control myogenesis. Mrf4 expression requires at least four elements, one of which may be shared with Myf5, providing a possible explanation for the linkage of these genes throughout vertebrate phylogeny. Further complexity is revealed by the demonstration that elements which control Mrf4 and Myf5 are embedded in an unrelated neighbouring gene.J. J. C. was supported by a Research Training Fellowship from the Medical Research Council (UK), which also paid for this work.Peer reviewe

Regulatory landscape of the vertebrate six2/six3 locus

Resumen del póster presentado al IX Meeting of the Spanish Society for Developmental Biology celebrado en Granada del 12 al 14 de noviembre de 2012.Peer Reviewe

CMOS First-Order All-Pass Filter With 2-Hz Pole Frequency

A CMOS fully integrated all-pass filter with an extremely low pole frequency of 2 Hz is introduced in this paper. It has 0.08-dB passband ripple and 0.029-mm 2 Si area. It has 0.38-mW power consumption in strong inversion with ±0.6-V power supplies. In subthreshold, it has 0.64-μW quiescent power and operates with ±200-mV dc supplies. Miller multiplication is used to obtain a large equivalent capacitor without excessive Si area. By varying the gain of the Miller amplifier, the pole frequency can be varied from 2 to 48 Hz. Experimental and simulation results of a test chip prototype in 130-nm CMOS technology validate the proposed circuit

Unravelling the function of Myf5 in the developing limb musculature

Motivation: Myogenesis is a complex process controlled by different networks, depending on the origin of different muscles, and that’s why there are different types of muscle distrophies. However, the myogenic cascade is always regulated by the same Myogenic Regulatory Factors (MRFs): Myf5, Mrf4, MyoD and MyoG. These transcription factors bind DNA and activate the expression of specific genes in particular progenitor cells that will give rise to the different muscles in the adult body. The MRFs’ cascade is initiated by Myf5, the first MRF to be expressed in the embryo. We and others have extensively studied the complex transcriptional regulation of the Myf5/Mrf4 locus using transgenic mice. Nowadays, we know that there are more than 25 regulatory elements controlling the expression of Mrf4 and Myf5 in a specific time frame and in particular embryonic progenitors.The limb enhancer is located 57 kb upstream of the Myf5 transcriptional start site. This enhancer controls Myf5 expression in limbs during development. While the mechanisms involved in the spatiotemporal regulation of Myf5 have been extensively studied at single-enhancer and global regulatory levels, the function of Myf5 in different subpopulations of muscle progenitor cells is still not clear. This project focuses on the characterization of a new allele in which the limb enhancer has been removed from the genome.Methods: To unravel the function of Myf5 in the developing limb musculature we have generated a new knock-out (KO) allele with CRISPR/Cas9 in which the limb enhancer has been targeted. Then, we have prepared RNA probes for In Situ Hybridisation (ISH) of genes that are expressed in limbs and are potential targets of Myf5 in other muscle progenitors (López-Mayorga et al., unpublished data), to test if the expression patters of these genes are modified in the KO allele. We are going to study three different embryonic stages: 10.5, 11.5 and 12.5 days post-coitum (dpc) by ISH using KO and wild type (WT) embryos. This time window was chosen to maximise the probability of detecting any pattern changes before the phenotype is rescued by the activity of MyoD, as previously shown. We are also preparing total RNA of fore-limbs from KO and WT embryos at 11.5 dpc to perform microarrays, which will give us some information about the genes regulated by Myf5 direct or indirectly. Finally, we will validate the results from microarrays by ISH and qPCR

±0.25-V Class-AB CMOS Capacitance Multiplier and Precision Rectifiers

Reduction of minimum supply requirements is a crucial aspect to decrease the power consumption in VLSI systems. A high-performance capacitance multiplier able to operate with supplies as low as ±0.25 V is presented. It is based on adaptively biased class-AB current mirrors which provide high current efficiency. Measurement results of a factor 11 capacitance multiplier fabricated in 180-nm CMOS technology verify theoretical claims. Moreover, low-voltage precision rectifiers based on the same class-AB current mirrors are designed and fabricated in the same CMOS process. They generate output currents over 100 times larger than the quiescent current. Both proposed circuits have 300-nW static power dissipation when operating with ±0.25-V supplies

An Op-Amp Approach for Bandpass VGAs With Constant Bandwidth

Two approaches to implement variable gain amplifiers based on Miller op-amps are discussed. One has true constant bandwidth while the other has essentially reduced bandwidth variations with varying gain. Servo-loops and ac coupling techniques with quasi floating gate transistors are used to provide a bandpass response with very low cutoff frequency in the range of hertz. In practice, one of the schemes is shown to have bandwidth variations close to a factor two while the second one has true constant bandwidth over the gain tuning range. Experimental results of test chip prototypes in 180-nm CMOS technology verify the theoretical claims

A polymorphic enhancer near GREM1 influences bowel cancer risk through differential CDX2 and TCF7L2 binding

Under a Creative Commons license.-- et al.A rare germline duplication upstream of the bone morphogenetic protein antagonist GREM1 causes aMendelian-dominant predisposition to colorectal cancer (CRC). The underlying disease mechanism is strong, ectopic GREM1 overexpression in the intestinal epithelium. Here, we confirm that a common GREM1 polymorphism, rs16969681, is also associated with CRC susceptibility, conferring ~20% differential risk in the general population. We hypothesized the underlying cause to be moderate differences inGREM1 expression. We showed that rs16969681 lies in a region of active chromatin with allele- and tissue-specific enhancer activity. The CRC high-risk allele was associated with stronger gene expression, and higher Grem1 mRNA levels increased the intestinal tumor burden in ApcMin mice. The intestine-specific transcription factor CDX2 and Wnt effector TCF7L2 bound near rs16969681, with significantly higher affinity for the risk allele, and CDX2 overexpression in CDX2/GREM1-negative cells caused re-expression of GREM1. rs16969681 influences CRC risk through effects on Wnt-driven GREM1 expression in colorectal tumors. © 2014 The Authors.Funding was provided from Cancer Research UK grant A/16459, an EU FP7 SYSCOL Consortium grant, and the EU COST colorectal cancer initiative. Core funding to the Wellcome Trust Centre for Human Genetics was provided from the Wellcome Trust (090532/Z/09/Z). J.L.G.-S. and J.J.C. were supported by the Spanish/FEDER government grants BFU2010-14839 and BFU2011-2292.Open Access funded by Wellcome Trust.Peer Reviewe

±0.3V Bulk-Driven Fully Differential Buffer with High Figures of Merit

A high performance bulk-driven rail-to-rail fully differential buffer operating from ±0.3V supplies in 180 nm CMOS technology is reported. It has a differential–difference input stage and common mode feedback circuits implemented with no-tail, high CMRR bulk-driven pseudo-differential cells. It operates in subthreshold, has infinite input impedance, low output impedance (1.4 kΩ), 86.77 dB DC open-loop gain, 172.91 kHz bandwidth and 0.684 μW static power dissipation with a 50-pF load capacitance. The buffer has power efficient class AB operation, a small signal figure of merit FOMSS = 12.69 MHzpFμW−1, a large signal figure of merit FOMLS = 34.89 (V/μs) pFμW−1, CMRR = 102 dB, PSRR+ = 109 dB, PSRR− = 100 dB, 1.1 μV/√Hz input noise spectral density, 0.3 mVrms input noise and 3.5 mV input DC offset voltage.Junta de Andalucía - Consejería de Economía, Conocimiento, Empresas y Universidades P18-FR-4317Agencia Estatal de Investigación - FEDER PID2019-107258RB-C3