Global gene expression during the human organogenesis: from transcription profiles to function predictions

Human embryogenesis includes an integrated set of complex yet coordinated development of different organs and tissues, which is regulated by the spatiotemporal expression of many genes. Deciphering the gene regulation profile is essential for understanding the molecular basis of human embryo development. While molecular and genetic studies in mouse have served as a valuable tool to understand mammalian development, significant differences exists in human and mouse development at morphological and genomic levels. Thus it is important to carry out research directly on human embryonic development. Here we will review some recent studies on gene regulation during human embryogenesis with particular focus on the period of organogenesis, which had not been well studied previously. We will highlight a gene expression database of human embryos from the 4(th) to the 9(th) week. The analysis of gene regulation during this period reveals that genes functioning in a given developmental process tend to be coordinately regulated during human embryogenesis. This feature allows us to use this database to identify new genes important for a particular developmental process/pathway and deduce the potential function of a novel gene during organogenesis. Such a gene expression atlas should serve as an important resource for molecular study of human development and pathogenesis.     

Nuclear transfer of embryonic cell nuclei to non-enucleated eggs in zebrafish, Danio rerio

We previously established a novel method for nuclear transfer in medaka (Oryzias latipes) using non-enucleated, diploidized eggs as recipients for adult somatic cell nuclei. Here we report the first attempt to apply this method to another fish species. To examine suitability of using non-enucleated eggs as recipients for nuclear transfer in the zebrafish (Danio rerio), we transferred blastula cell nuclei from a wild-type donor strain to non-enucleated, unfertilized eggs from a golden recipient strain. As a result, 31 of 184 (16.8%) operated eggs developed normally and reached the adult stage. Twenty-eight (15.2%) of these transplants showed wild-type phenotype and the remaining three (1.6%) were golden. Except for one individual that exhibited diploid/tetraploid mosaicism, all of the wild-type nuclear transplants were either triploid or diploid. While all of 19 triploid transplants were infertile, a total of six transplants (21.4%) were fertile (five of the eight diploid transplants and one transplant exhibiting ploidy mosaicism). Except for one diploid individual, all of the fertile transplants transferred both the wild-type golden gene allele (slc24a5) as well as the phenotype, the wild-type body color, to their F(1) and F(2) progeny in a typical Mendelian fashion. PCR analysis of slc24a5 suggested that triploidy originated from a fused nucleus in the diploid donor and haploid recipient nuclei, and that the sole origin of diploidy was the diploid donor nucleus. The results of the present study demonstrated the suitability of using non-enucleated eggs as recipients for nuclear transfer experiments in zebrafish.     

Cortical network from human embryonic stem cells

The connection of embryonic stem cell technology and developmental biology provides valuable tools to decipher the mechanisms underlying human brain development and diseases, especially among neuronal populations, that are not readily available in primary cultures. It is obviously the case of neurons forming the human cerebral cortex. In the images that are presented, the neurons were generated in vitro from human embryonic stem cells via forebrain-like progenitors. Maintained in culture for prolonged time, they acquired a mainly glutamatergic phenotype and morphological characteristics of cortical pyramidal neurons, including dendritic spines, and formed spectacular networks.     

Evaluation of differential antitermitic activities of Lantana camara oven‐dried tissues against Reticulitermes virginicus (Isoptera: Rhinotermitidae)

Abstract Chemical‐treated soil or physical barriers have been the most commonly used approach for termite management. We hypothesized that a barrier of soil incorporated with oven‐dried Lantana camara L. tissues could prevent termite infestation. We first examined the antitermitic effects of the dried tissues from two cultivars (‘Mozelle’ and ‘New Gold’) on the subterranean termite, Reticulitermes virginicus (Banks) (Isoptera: Rhinotermitidae). Results show that all of the tissues of Mozelle had greater antitermic activity than corresponding tissues of New Gold, and leaves had greater termiticidal effects than flowers and stems. When termites were exposed to the test materials in a no‐choice bioassay, the 24‐day test resulted in a significant reduction of survival (52.5%–88.6%), running speed (18.2%–37.3%), live weight (21.8%–53.5%) and body water content (33.2%–56.2%) compared to the control. The consumption of leaves and flowers was exiguous. When used as 25% tissue mulch‐barrier, the oven‐dried lantana tissues decreased termite tunneling and wood consumption and increased termite mortality. The decreased survival, vigor, and low consumption indicate a toxic and anti‐feeding property of the materials tested. The results therefore support our hypothesis that the dried lantana tissues possess antitermitic activities.     

New glimpses of caveolin-1 functions in embryonic development and human diseases

Caveolin-1 (Cav-1) isoforms, including Cav-1α and Cav-1β, were identified as integral membrane proteins and the major components of caveolae. Cav-1 proteins are highly conserved during evolution from {itCaenorhabditis elegans} to human and are capable of interacting with many signaling molecules through their caveolin scaffolding domains to regulate the activities of multiple signaling pathways. Thus, Cav-1 plays crucial roles in the regulation of cellular proliferation, differentiation and apoptosis in a cell-specific and contextual manner. In addition, Cav-1 is essential for embryonic development of vertebrates owing to its regulation of BMP, Wnt, TGF-β and other key signaling molecules. Moreover, Cav-1 is mainly expressed in terminally differentiated cells and its abnormal expression is often associated with human diseases, such as tumor progression, cardiovascular diseases, fibrosis, lung regeneration, and diseases related to virus. In this review, we will further discuss the potential of Cav-1 as a target for disease therapy and multiple drug resistance.     

Efficient derivation of human cardiac precursors and cardiomyocytes from pluripotent human embryonic stem cells with small molecule induction

To date, the lack of a suitable human cardiac cell source has been the major setback in regenerating the human myocardium, either by cell-based transplantation or by cardiac tissue engineering. Cardiomyocytes become terminally-differentiated soon after birth and lose their ability to proliferate. There is no evidence that stem/progenitor cells derived from other sources, such as the bone marrow or the cord blood, are able to give rise to the contractile heart muscle cells following transplantation into the heart. The need to regenerate or repair the damaged heart muscle has not been met by adult stem cell therapy, either endogenous or via cell delivery. The genetically stable human embryonic stem cells (hESCs) have unlimited expansion ability and unrestricted plasticity, proffering a pluripotent reservoir for in vitro derivation of large supplies of human somatic cells that are restricted to the lineage in need of repair and regeneration. Due to the prevalence of cardiovascular disease worldwide and acute shortage of donor organs, there is intense interest in developing hESC-based therapies as an alternative approach. However, how to channel the wide differentiation potential of pluripotent hESCs efficiently and predictably to a desired phenotype has been a major challenge for both developmental study and clinical translation. Conventional approaches rely on multi-lineage inclination of pluripotent cells through spontaneous germ layer differentiation, resulting in inefficient and uncontrollable lineage-commitment that is often followed by phenotypic heterogeneity and instability, hence, a high risk of tumorigenicity (see a schematic in Fig. 1A). In addition, undefined foreign/animal biological supplements and/or feeders that have typically been used for the isolation, expansion, and differentiation of hESCs may make direct use of such cell-specialized grafts in patients problematic. To overcome these obstacles, we have resolved the elements of a defined culture system necessary and sufficient for sustaining the epiblast pluripotence of hESCs, serving as a platform for de novo derivation of clinically-suitable hESCs and effectively directing such hESCs uniformly towards clinically-relevant lineages by small molecules (see a schematic in Fig. 1B). After screening a variety of small molecules and growth factors, we found that such defined conditions rendered nicotinamide (NAM) sufficient to induce the specification of cardiomesoderm direct from pluripotent hESCs that further progressed to cardioblasts that generated human beating cardiomyocytes with high efficiency (Fig. 2). We defined conditions for induction of cardioblasts direct from pluripotent hESCs without an intervening multi-lineage embryoid body stage, enabling well-controlled efficient derivation of a large supply of human cardiac cells across the spectrum of developmental stages for cell-based therapeutics.     

Chorein,the protein responsible for chorea-acanthocytosis,interacts with β-adducin and β-actin

Chorea-acanthocytosis (ChAc) is an autosomal, recessive hereditary disease characterized by striatal neurodegeneration and acanthocytosis, and caused by loss of function mutations in the vacuolar protein sorting 13 homolog A (VPS13A) gene. VPS13A encodes chorein whose physiological function at the molecular level is poorly understood. In this study, we show that chorein interacts with β-adducin and β-actin. We first compare protein expression in human erythrocyte membranes using proteomic analysis. Protein levels of β-adducin isoform 1 and β-actin are markedly decreased in erythrocyte membranes from a ChAc patient. Subsequent co-immunoprecipitation (co-IP) and reverse co-IP assays using extracts from chorein-overexpressing human embryonic kidney 293 (HEK293) cells, shows that β-adducin (isoforms 1 and 2) and β-actin interact with chorein. Immunocytochemical analysis using chorein-overexpressing HEK293 cells demonstrates co-localization of chorein with β-adducin and β-actin. In addition, immunoreactivity of β-adducin isoform 1 is significantly decreased in the striatum of gene-targeted ChAc-model mice. Adducin and actin are membrane cytoskeletal proteins, involved in synaptic function. Expression of β-adducin is restricted to the brain and hematopoietic tissues, corresponding to the main pathological lesions of ChAc, and thereby implicating β-adducin and β-actin in ChAc pathogenesis.     

The Antiviral Activities of ISG15

Post-translational protein modification is an important strategy for the regulation of the cell proteome independent of the need for new gene expression. Ubiquitin and ubiquitin-like modifiers mediate the regulation of protein levels, signaling pathways, vesicular trafficking, and many other cellular processes through their covalent conjugation to proteins. Interferon stimulated gene 15 (ISG15) is a ubiquitin-like modifier induced by type I interferon. In addition to conjugating to potentially hundreds of target proteins, ISG15 can be found in an unconjugated form both inside of the cell and released from interferon stimulated cells into the extracellular environment. Due to its robust expression after type I interferon stimulation and the broad panel of proteins that it targets, ISG15 has drawn much attention as a potential regulator of the immune response and has been shown to mediate protection in a number of different viral infection models. Here we will review the current state of the field of ISG15, the viruses against which ISG15 mediates protection, and the mechanisms by which ISG15 exerts antiviral activity.