Abstract. Mice with a targeted mutation in the myogenic basic helix-loop-helix regulatory protein myogenin have severe muscle defects resulting in perinatal death. In this report, the effect of myogenin's absence on embryonic and fetal development is investigated. The initial events of somite differentiation occurred normally in the myogenin-mutant embryos. During primary myogenesis, muscle masses in mutant embryos developed simultaneously with control siblings, although muscle differentiation within the mutant muscle masses was delayed. More dramatic effects were observed when secondary myofibers form. During this time, very little muscle formation took place in the mutants, suggesting that the absence of myogenin affected secondary myogenesis more severely than primary myogenesis. Monitoring mutant neonates with fiber type-specific myosin isoforms indicated that different fiber types were present in the residual muscle. No evidence was found to indicate that myogenin was required for the formation of muscle in one region of the embryo and not another. The expression patterns of a MyoD-lacZ transgene in myogenin-mutant embryos demonstrated that myogenin was not essential for the activation of the MyoD gene. Together, these results indicate that late stages of embryogenesis are more dependent on myogenin than early stages, and that myogenin is not required for the initial aspects of myogenesis, including myotome formation and the appearance of myoblasts. SKELETAL muscle in vertebrates originates from somitic mesoderm as pluripotent mesodermal cells become committed to a myogenic fate. Committed myoblasts populate areas throughout the developing embryo, ultimately differentiating into bundles of multinucleate myofibers. Four key players in myogenic events are the basic helix-loop-helix (bHLH) ~ regulatory proteins: MyoD, Myf5, myogenin, and MRF4 (for recent reviews see Emerson, 1993;Weintraub, 1993;Olson and Klein, 1994). These muscle-specific transcription factors are individually able to initiate the entire muscle differentiation program when introduced into tissue culture cells of nonmuscle origin. Using gene-knockout technology, several laboratories have created mice lacking functional myogenic bHLH factors and are now providing useful models for studying skeletal muscle development (Braun et al

In the preimplantation mouse embryo, TEAD4 is critical to establishing the trophectoderm (TE)-specific transcriptional program and segregating TE from the inner cell mass (ICM). However, TEAD4 is expressed in the TE and the ICM. Thus, differential function of TEAD4 rather than expression itself regulates specification of the first two cell lineages. We used ChIP sequencing to define genomewide TEAD4 target genes and asked how transcription of TEAD4 target genes is specifically maintained in the TE. Our analyses revealed an evolutionarily conserved mechanism, in which lack of nuclear localization of TEAD4 impairs the TE-specific transcriptional program in inner blastomeres, thereby allowing their maturation toward the ICM lineage. Restoration of TEAD4 nuclear localization maintains the TE-specific transcriptional program in the inner blastomeres and prevents segregation of the TE and ICM lineages and blastocyst formation. We propose that altered subcellular localization of TEAD4 in blastomeres dictates first mammalian cell fate specification.A llocation of blastomeres to outside and inside positions during preimplantation mammalian development initiates specification of the first two cell lineages, the trophectoderm (TE) and the inner cell mass (ICM) (1, 2). Successful progression of TE and ICM fate specification and proper development of the preimplantation embryo depends on differential transcriptional programs that are instigated and maintained within the outer and inner cells. Gene-KO studies in mice showed TEAD4 as the master orchestrator of the TE-specific transcriptional program (3-5). TEAD4-null embryos do not mature to the blastocyst stage and TEAD4-null blastomeres lack expression of TE-specific master regulators like CDX2, GATA3, and EOMES (3, 4). However, they maintain expression of ICM-specific factors like OCT4 and NANOG.Interestingly, TEAD4 expression is maintained both in cells of TE and ICM lineages, as well as in the TE-derived trophoblast stem cells (TSCs) and ICM-derived ES cells (ESCs) (5, 6). Thus, questions are raised as to how TEAD4 selectively orchestrates the TE/TSC-specific transcriptional program but not the ICM/ ESC-specific transcriptional program. The current model predicts that the presence vs. the absence of a TEAD4 cofactor, yesassociated protein (YAP), modulates TEAD4 function at its target genes in outer vs. inner blastomeres (6), leading to the segregation of the TE and ICM lineages. However, YAP-null mouse embryos do not show preimplantation developmental defects (7), indicating that, unlike TEAD4, YAP function is dispensable during TE and ICM fate determination. It is proposed that another YAP-related cofactor, WWTR1 (i.e., TAZ), could compensate for the absence of YAP during early development (6). However, the mode of TAZ function during TE and ICM specification is unknown. Furthermore, direct targets of TEAD4 have not been identified in the TE or in trophoblast cells. Thus, definitive experiments have not been performed to conclude that loss of cofactor function/recruitmen...

Histone methylation is an important regulator of gene expression; its coordinated activity is critical in complex developmental processes such as hematopoiesis. Disruptor of telomere silencing 1-like (DOT1L) is a unique histone methyltransferase that specifically methylates histone H3 at lysine 79. We analyzed Dot1L-mutant mice to determine influence of this enzyme on embryonic hematopoiesis. Mutant mice developed more slowly than wild-type embryos and died between embryonic days 10.5 and 13.5, displaying a striking anemia, especially apparent in small vessels of the yolk sac. Further, a severe, selective defect in erythroid, but not myeloid, differentiation was observed. Erythroid progenitors failed to develop normally, showing retarded progression through the cell cycle, accumulation during G 0 /G 1 stage, and marked increase in apoptosis in response to erythroid growth factors. GATA2, a factor essential for early erythropoiesis, was significantly reduced in Dot1L-deficient cells, whereas expression of PU.1, a transcription factor that inhibits erythropoiesis and promotes myelopoiesis, was increased. These data suggest a model whereby DOT1L-dependent lysine 79 of histone H3 methylation serves as a critical regulator of a differentiation switch during early hematopoiesis, regulating steady-state levels of GATA2 and PU.1 transcription, thus controlling numbers of circulating erythroid and myeloid cells. IntroductionAmong the first differentiated cell types to emerge in the developing mammalian embryo are the blood cells. In the mouse, the process of blood development, hematopoiesis, begins at approximately embryonic day 7.0-7.5 (E7.0-E7.5), when cells originating in the primitive streak migrate to the site of yolk sac formation. 1 By E7.5, the cells coalesce into blood islands, where they mature, proliferate, and differentiate. 2 These early hematopoietic progenitors, termed primitive erythroid colony-forming cells, are nucleated red cells, which express primitive globins and can carry oxygen to nourish the developing embryo on the initiation of blood flow after E8.5. 1,[3][4][5] The presence of these primitive progenitors is transient, peaking in numbers at E8.0 and disappearing by E9.0, 2 whereas the progeny erythrocytes persist throughout gestation. 6 After E8.5, a second wave of hematopoietic progenitors emerges from a variety of sites, including the vasculature about the aorta-gonadmesonephros and the yolk sac. These cells enter the circulation and migrate to the developing fetal liver. There, they proliferate and undergo "definitive" maturation, giving rise to multiple adult hematopoietic lineages, including mature, enucleated erythrocytes. 7 This multistep process of hematopoiesis and the fate decisions of the developing cells are regulated by the precisely controlled, sequential induction and silencing of gene expression in response to a variety of growth and differentiation factors. 8 The identity of the cell-type specific genes that direct differentiation, the factors controlling their expression, and...

Because of their deleterious effects on developing organisms, ribosomal protein (RP) mutations have been poorly described in mammals, and only a few heterozygous mutations have been shown to be viable. This observation is believed to be due to the fact that each RP is an essential component in the assembly of a functional stable ribosome. Here, we created gene targeted mutant mice lacking HIP/RPL29, an RP associated with translationally active ribosomes in eukaryotes. In contrast to other RP mutants, HIP/RPL29 null mice are viable but are up to 50% smaller than their control littermates at weaning age. In null embryos, delayed global growth is first observed around mid-gestation, and postnatal lethality due to low birth weight results in distortion of the Mendelian ratio. Prenatal growth defects are not fully compensated for during adulthood, and null animals display proportionately smaller organs and stature, and reach sexual maturity considerably later when compared with their control siblings. Additionally, HIP/RPL29 null embryonic fibroblasts have decreased rates of proliferation and protein synthesis and exhibit reduced steady state levels of core RPs. Altogether, our findings provide conclusive genetic evidence that HIP/RPL29 functions as an important regulator of global growth by modulating the rate of protein synthesis. Developmental Dynamics 236:447-460, 2007.