Integrated Functions of Pax3 and Pax7 in the Regulation of Proliferation, Cell Size and Myogenic Differentiation

Pax3 and Pax7 are paired-box transcription factors with roles in developmental and adult regenerative myogenesis. Pax3 and Pax7 are expressed by postnatal satellite cells or their progeny but are down regulated during myogenic differentiation. We now show that constitutive expression of Pax3 or Pax7 in either satellite cells or C2C12 myoblasts results in an increased proliferative rate and decreased cell size. Conversely, expression of dominant-negative constructs leads to slowing of cell division, a dramatic increase in cell size and altered morphology. Similarly to the effects of Pax7, retroviral expression of Pax3 increases levels of Myf5 mRNA and MyoD protein, but does not result in sustained inhibition of myogenic differentiation. However, expression of Pax3 or Pax7 dominant-negative constructs inhibits expression of Myf5, MyoD and myogenin, and prevents differentiation from proceeding. In fibroblasts, expression of Pax3 or Pax7, or dominant-negative inhibition of these factors, reproduce the effects on cell size, morphology and proliferation seen in myoblasts. Our results show that in muscle progenitor cells, Pax3 and Pax7 function to maintain expression of myogenic regulatory factors, and promote population expansion, but are also required for myogenic differentiation to proceed

Expression Profiling of FSHD-1 and FSHD-2 Cells during Myogenic Differentiation Evidences Common and Distinctive Gene Dysregulation Patterns

BACKGROUND: Determine global gene dysregulation affecting 4q-linked (FSHD-1) and non 4q-linked (FSHD-2) cells during early stages of myogenic differentiation. This approach has been never applied to FSHD pathogenesis. METHODOLOGY/PRINCIPAL FINDINGS: By in vitro differentiation of FSHD-1 and FSHD-2 myoblasts and gene chip analysis we derived that gene expression profile is altered only in FSHD-1 myoblasts and FSHD-2 myotubes. The changes seen in FSHD-1 regarded a general defect in cell cycle progression, probably due to the upregulation of myogenic markers PAX3 and MYOD1, and a deficit of factors (SUV39H1 and HMGB2) involved in D4Z4 chromatin conformation. On the other hand, FSHD-2 mytubes were characterized by a general defect in RNA metabolism, protein synthesis and degradation and, to a lesser extent, in cell cycle. Common dysregulations regarded genes involved in response to oxidative stress and in sterol biosynthetic process. Interestingly, our results also suggest that miRNAs might be implied in both FSHD-1 and FSHD-2 gene dysregulation. Finally, in both cell differentiation systems, we did not observe a gradient of altered gene expression throughout the 4q35 chromosome. CONCLUSIONS/SIGNIFICANCE: FSHD-1 and FSHD-2 cells showed, in different steps of myogenic differentiation, a global deregulation of gene expression rather than an alteration of expression of 4q35 specific genes. In general, FSHD-1 and FSHD-2 global gene deregulation interested common and distinctive biological processes. In this regard, defects of cell cycle progression (FSHD-1 and to a lesser extent FSHD-2), protein synthesis and degradation (FSHD-2), response to oxidative stress (FSHD-1 and FSHD-2), and cholesterol homeostasis (FSHD-1 and FSHD-2) may in general impair a correct myogenesis. Taken together our results recapitulate previously reported defects of FSHD-1, and add new insights into the gene deregulation characterizing both FSHD-1 and FSHD-2, in which miRNAs may play a role

Segregation of myoblast fusion and muscle-specific gene expression by distinct ligand-dependent inactivation of GSK-3β

Myogenic differentiation involves myoblast fusion and induction of muscle-specific gene expression, which are both stimulated by pharmacological (LiCl), genetic, or IGF-I-mediated GSK-3β inactivation. To assess whether stimulation of myogenic differentiation is common to ligand-mediated GSK-3β inactivation, myoblast fusion and muscle-specific gene expression were investigated in response to Wnt-3a. Moreover, crosstalk between IGF-I/GSK-3β/NFATc3 and Wnt/GSK-3β/β-catenin signaling was assessed. While both Wnt-3a and LiCl promoted myoblast fusion, muscle-specific gene expression was increased by LiCl, but not by Wnt-3a or β-catenin over-expression. Furthermore, LiCl and IGF-I, but not Wnt-3a, increased NFATc3 transcriptional activity. In contrast, β-catenin-dependent transcriptional activity was increased by Wnt-3a and LiCl, but not IGF-I. These results for the first time reveal a segregated regulation of myoblast fusion and muscle-specific gene expression following stimulation of myogenic differentiation in response to distinct ligand-specific signaling routes of GSK-3β inactivation

Comparative Analyses by Sequencing of Transcriptomes during Skeletal Muscle Development between Pig Breeds Differing in Muscle Growth Rate and Fatness

Understanding the dynamics of muscle transcriptome during development and between breeds differing in muscle growth is necessary to uncover the complex mechanism underlying muscle development. Herein, we present the first transcriptome-wide longissimus dorsi muscle development research concerning Lantang (LT, obese) and Landrace (LR, lean) pig breeds during 10 time-points from 35 days-post-coitus (dpc) to 180 days-post-natum (dpn) using Solexa/Illumina's Genome Analyzer. The data demonstrated that myogenesis was almost completed before 77 dpc, but the muscle phenotypes were still changed from 77 dpc to 28 dpn. Comparative analysis of the two breeds suggested that myogenesis started earlier but progressed more slowly in LT than in LR, the stages ranging from 49 dpc to 77 dpc are critical for formation of different muscle phenotypes. 595 differentially expressed myogenesis genes were identified, and their roles in myogenesis were discussed. Furthermore, GSK3B, IKBKB, ACVR1, ITGA and STMN1 might contribute to later myogenesis and more muscle fibers in LR than LT. Some myogenesis inhibitors (ID1, ID2, CABIN1, MSTN, SMAD4, CTNNA1, NOTCH2, GPC3 and HMOX1) were higher expressed in LT than in LR, which might contribute to more slow muscle differentiation in LT than in LR. We also identified several genes which might contribute to intramuscular adipose differentiation. Most important, we further proposed a novel model in which MyoD and MEF2A controls the balance between intramuscular adipogenesis and myogenesis by regulating CEBP family; Myf5 and MEF2C are essential during the whole myogenesis process while MEF2D affects muscle growth and maturation. The MRFs and MEF2 families are also critical for the phenotypic differences between the two pig breeds. Overall, this study contributes to elucidating the mechanism underlying muscle development, which could provide valuable information for pig meat quality improvement

Motives and comprehension in a public goods game with induced emotions

This study analyses the sensitivity of public goods contributions through the lens of psychological motives. We report the results of a public goods experiment in which subjects were induced with the motives of care and anger through autobiographical recall. Subjects' preferences, beliefs, and perceptions under each motive are compared with those of subjects experiencing a neutral autobiographical recall control condition. We find, but only for those subjects with the highest comprehension of the game, that care elicits significantly higher contributions than anger, with the control treatment in between. This positive influence of the care motive on unconditional giving is accounted for partly by preferences for giving and partly by the beliefs concerning greater contributions by others. Anger also affects attention to own and other's payoffs (using mouse tracking) and perceptions of the game's incentive structure (cooperative or competitive)

Mouse models of breast cancer metastasis

Metastatic spread of cancer cells is the main cause of death of breast cancer patients, and elucidation of the molecular mechanisms underlying this process is a major focus in cancer research. The identification of appropriate therapeutic targets and proof-of-concept experimentation involves an increasing number of experimental mouse models, including spontaneous and chemically induced carcinogenesis, tumor transplantation, and transgenic and/or knockout mice. Here we give a progress report on how mouse models have contributed to our understanding of the molecular processes underlying breast cancer metastasis and on how such experimentation can open new avenues to the development of innovative cancer therapy

Key signaling nodes in mammary gland development and cancer: β-catenin

β-Catenin plays important roles in mammary development and tumorigenesis through its functions in cell adhesion, signal transduction and regulation of cell-context-specific gene expression. Studies in mice have highlighted the critical role of β-catenin signaling for stem cell biology at multiple stages of mammary development. Deregulated β-catenin signaling disturbs stem and progenitor cell dynamics and induces mammary tumors in mice. Recent data showing deregulated β-catenin signaling in metaplastic and basal-type tumors suggest a similar link to reactivated developmental pathways and human breast cancer. The present review will discuss β-catenin as a central transducer of numerous signaling pathways and its role in mammary development and breast cancer

Increased throughput and ultra-high mass resolution in DESI FT-ICR MS imaging through new-generation external data acquisition system and advanced data processing approaches

Desorption electrospray ionisation-mass spectrometry imaging (DESI-MSI) is a powerful imaging technique for the analysis of complex surfaces. However, the often highly complex nature of biological samples is particularly challenging for MSI approaches, as options to appropriately address mass spectral complexity are limited. Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) offers superior mass accuracy and mass resolving power, but its moderate throughput inhibits broader application. Here we demonstrate the dramatic gains in mass resolution and/or throughput of DESI-MSI on an FT-ICR MS by developing and implementing a sophisticated data acquisition and data processing pipeline. The presented pipeline integrates, for the first time, parallel ion accumulation and detection, post-processing absorption mode Fourier transform and pixel-by-pixel internal re-calibration. To achieve that, first, we developed and coupled an external high-performance data acquisition system to an FT-ICR MS instrument to record the time-domain signals (transients) in parallel with the instrument’s built-in electronics. The recorded transients were then processed by the in-house developed computationally-efficient data processing and data analysis software. Importantly, the described pipeline is shown to be applicable even to extremely large, up to 1 TB, imaging datasets. Overall, this approach provides improved analytical figures of merits such as: (i) enhanced mass resolution at no cost in experimental time; and (ii) up to 4-fold higher throughput while maintaining a constant mass resolution. Using this approach, we not only demonstrate the record 1 million mass resolution for lipid imaging from brain tissue, but explicitly demonstrate such mass resolution is required to resolve the complexity of the lipidome