Transmembrane and intracellular signal transduction during palatal ontogeny.

Investigations of the pathogenesis of cleft palate have been hampered by the complexity of normal craniofacial ontogenesis and by an incomplete understanding of mechanisms involved in normal palate development. This article deals with cellular aspects of the developing craniofacial region and discusses the facts and problems in the field as they relate to the normal development of the secondary palate, with emphasis on the regulation of hormone- and growth factor-induced transmembrane signalling and intracellular signal transduction.     

Synthetic biology of minimal systems

“Synthetic biology” is a concept that has developed together with, or slightly after, “systems biology”. But while systems biology aims at the full understanding of large systems by integrating more and more details into their models, synthetic biology phrases different questions, namely: what particular biological function could be obtained with a certain known subsystem of reduced complexity; can this function be manipulated or engineered in artificial environments or genetically modified organisms; and if so, how? The most prominent representation of synthetic biology has so far been microbial engineering by recombinant DNA technology, employing modular concepts known from information technology. However, there are an increasing number of biophysical groups who follow similar strategies of dissecting cellular processes and networks, trying to identify functional minimal modules that could then be combined in a bottom-up approach towards biology. These modules are so far not as particularly defined by their impact on DNA processing, but rather influenced by core fields of biophysics, such as cell mechanics and membrane dynamics. This review will give an overview of some classical and some quite new biophysical strategies for constructing minimal systems of certain cellular modules. We will show that with recent advances in understanding of cytoskeletal and membrane elements, the time might have come to experimentally challenge the concept of a minimal cell.     

Expression of T1Rs and gustducin in palatal taste buds of mice

The palatal region of the oral cavity in rodents houses 100-300 taste buds and is particularly sensitive to sweet and umami compounds; yet, few studies have examined the expression patterns of transduction-related molecules in this taste field. We investigated the interrelationships between members of the T1R family and between each T1R and gustducin in palatal taste buds. Similar to lingual taste buds, T1R1 and T1R2 are generally expressed in separate palatal taste cells. In contrast to lingual taste buds, however, T1R2 and T1R3-positive palatal taste cells almost always coexpress gustducin, suggesting that sweet taste transduction in the palate is almost entirely dependent on gustducin. T1R1-positive palate taste cells coexpress gustducin about half the time, suggesting that other G proteins may contribute to the transduction of umami stimuli in this taste field.     

Bridging the gaps in systems biology

Systems biology aims at creating mathematical models, i.e., computational reconstructions of biological systems and processes that will result in a new level of understanding—the elucidation of the basic and presumably conserved “design” and “engineering” principles of biomolecular systems. Thus, systems biology will move biology from a phenomenological to a predictive science. Mathematical modeling of biological networks and processes has already greatly improved our understanding of many cellular processes. However, given the massive amount of qualitative and quantitative data currently produced and number of burning questions in health care and biotechnology needed to be solved is still in its early phases. The field requires novel approaches for abstraction, for modeling bioprocesses that follow different biochemical and biophysical rules, and for combining different modules into larger models that still allow realistic simulation with the computational power available today. We have identified and discussed currently most prominent problems in systems biology: (1) how to bridge different scales of modeling abstraction, (2) how to bridge the gap between topological and mechanistic modeling, and (3) how to bridge the wet and dry laboratory gap. The future success of systems biology largely depends on bridging the recognized gaps.     

Overexpression of iNOS and down-regulation of BMPs-2, 4 and 7 in retinoic acid induced cleft palate formation

The present work studied the induction of cleft palate formation in embryos developed from pregnant BALB/c mice treated orally with retinoic acid (RA). Previous studies on mature somatic cell types showed that RA exerted inhibitory effects on inducible nitric oxide synthase (iNOS) production. For the first time, our study has shown that RA actually stimulates significant expression of iNOS at specific zones of the affected embryonic palatal tissues at three consecutive stages, from gestation day 13 (GD13) to day 16 (GD16). Enzymatically, iNOS facilitates intracellular nitric oxide (NO) synthesis from L-arginine. When NO reacts with reactive superoxides it may result in irreparable cell injury. NO was also reported to induce apoptosis in some mammalian cell systems. Based on our findings, we propose that such an increase in NO production might be associated with apoptosis in the embryonic palatal tissues in the RA-treated mice. The detrimental effects of NO resulted in a reduction in proliferating palatal cells and therefore disturbed the normal plasticity of the palatal shelves. With iNOS overexpression, our findings also showed that there was significant concomitant down-regulation in the expressions of Bone Morphogenetic Proteins (BMPs) -2, 4, and 7 with regional variations particularly in the palatal mesenchymal cells for those embryos developing cleft palate. Since specific spatial and temporal expressions of BMPs -2, 4, and 7 are critical during normal palatal morphogenesis, any deficiency in the epithelial-mesenchymal interaction may result in retarding growth at the embryonic palatal shelves. Taken together, our study has demonstrated cleft palate formation in the BALB/c embryos involved overexpression of iNOS and down-regulation of BMPs-2, 4 and 7.     

Advantages and clinical applications of natural killer cells in cancer immunotherapy

The past decade has witnessed a burgeoning of research and further insight into the biology and clinical applications of natural killer (NK) cells. Once thought to be simple innate cells important only as cytotoxic effector cells, our understanding of NK cells has grown to include memory-like responses, the guidance of adaptive responses, tissue repair, and a delicate paradigm for how NK cells become activated now termed “licensing” or “arming.” Although these cells were initially discovered and named for their spontaneous ability to kill tumor cells, manipulating NK cells in therapeutic settings has proved difficult and complex in part due to our emerging understanding of their biology. Therapies involving NK cells may either activate endogenous NK cells or involve transfers of exogenous cells by hematopoietic stem cell transplantation or adoptive cell therapy. Here, we review the basic biology of NK cells, highlighting characteristics which make NK cells particularly useful in cancer therapies. We also explore current treatment strategies that have been used for cancer as well as discuss potential future directions for the field.     

Mechanisms of bacterial morphogenesis: Evolutionary cell biology approaches provide new insights

How Darwin's “endless forms most beautiful” have evolved remains one of the most exciting questions in biology. The significant variety of bacterial shapes is most likely due to the specific advantages they confer with respect to the diverse environments they occupy. While our understanding of the mechanisms generating relatively simple shapes has improved tremendously in the last few years, the molecular mechanisms underlying the generation of complex shapes and the evolution of shape diversity are largely unknown. The emerging field of bacterial evolutionary cell biology provides a novel strategy to answer this question in a comparative phylogenetic framework. This relatively novel approach provides hypotheses and insights into cell biological mechanisms, such as morphogenesis, and their evolution that would have been difficult to obtain by studying only model organisms. We discuss the necessary steps, challenges, and impact of integrating “evolutionary thinking” into bacterial cell biology in the genomic era.     

Intracellular dynamics of Smad-mediated TGFbeta signaling

The transforming growth factor-beta (TGFbeta) family represents a class of signaling molecules that plays a central role in morphogenesis, growth, and cell differentiation during normal embryonic development. Members of this growth factor family are particularly vital to development of the mammalian secondary palate where they regulate palate mesenchymal cell proliferation and extracellular matrix synthesis. Such regulation is particularly critical since perturbation of either cellular process results in a cleft of the palate. While the cellular and phenotypic effects of TGFbeta on embryonic craniofacial tissue have been extensively catalogued, the specific genes that function as downstream mediators of TGFbeta action in the embryo during palatal ontogenesis are poorly defined. Embryonic palatal tissue in vivo and murine embryonic palate mesenchymal (MEPM) cells in vitro secrete and respond to TGFbeta. In the current study, elements of the Smad component of the TGFbeta intracellular signaling system were identified and characterized in cells of the embryonic palate and functional activation of the Smad pathway by TGFbeta1, TGFbeta2, and TGFbeta3 was demonstrated. TGFbeta-initiated Smad signaling in cells of the embryonic palate was found to result in: (1) phosphorylation of Smad 2; (2) nuclear translocation of the Smads 2, 3, and 4 protein complex; (3) binding of Smads 3 and 4 to a consensus Smad binding element (SBE) oligonucleotide; (4) transactivation of transfected reporter constructs, containing TGFbeta-inducible Smad response elements; and (4) increased expression of gelatinases A and B (endogenous genes containing Smad response elements) whose expression is critical to matrix remodeling during palatal ontogenesis. Collectively, these data point to the presence of a functional Smad-mediated TGFbeta signaling system in cells of the developing murine palate.     

Ultrastructural evidence of contractile systems in mouse palates prior to rotation.

Previous studies have shown that the palatal shelves of mouse embryos synthesize the contractile proteins actin and myosin at a rate equal to that of tongue just prior to shelf movement (day 14.5). The purpose of this study was to examine the morphology of the palatal shelves for evidence of a contractile system. Myosin ATPase histochemistry was performed on frozen sections of day-14.5 fetal mouse heads. Three areas of the palatal shelves gave a positive reaction: 1) A reaction product typical of skeletal muscle on the oral side of the posterior palate (region 1); 2) a “heavy-diffuse” reaction product on the tongue side extending from the top mid-palate to the posterior end (region 2); and 3) a “light-diffuse” reaction product along the oral epithelium in the mid-palate (region 3). Electron microscopy of excised day-14.5 palates was carried out after fixation in glutaraldehyde or an acrolein-dichromate solution. Region 1 contained a large area of developing and adultlike skeletal muscle. In the area of region 2 a large population of filamentous-rich mesenchymal cells was observed. In addition, large neurons coursing through both contractile systems were noted. Preliminary observations in region 3 indicated the possibility of a primitive (nonmuscle) contractile system in that area. The contractile and nervous systems in the palate, prior to rotation, indicate the possibility that an innervated embryonic muscle system may provide the “intrinsic shelf force” to rotate the shelves.     

Meeting report: Signal transduction meets systems biology

ABSTRACT: In the 21st century, systems-wide analyses of biological processes are getting more and more realistic. Especially for the in depth analysis of signal transduction pathways and networks, various approaches of systems biology are now successfully used. The EU FP7 large integrated project SYBILLA (Systems Biology of T-cell Activation in Health and Disease) coordinates such an endeavor. By using a combination of experimental data sets and computational modelling, the consortium strives for gaining a detailed and mechanistic understanding of signal transduction processes that govern T-cell activation. In order to foster the interaction between systems biologists and experimentally working groups, SYBILLA co-organized the 15th meeting "Signal Transduction: Receptors, Mediators and Genes" together with the Signal Transduction Society (STS). Thus, the annual STS conference, held from November 7 to 9, 2011 in Weimar, Germany, provided an interdisciplinary forum for research on signal transduction with a major focus on systems biology addressing signalling events in T-cells. Here we report on a selection of ongoing projects of SYBILLA and how they were discussed at this interdisciplinary conference.     

Programmed longevity,youthspan, and juventology

The identification of conserved genes and pathways that regulate lifespan but also healthspan has resulted in an improved understanding of the link between nutrients, signal transduction proteins, and aging but has also provided evidence for the existence of multiple “longevity programs,” which are selected based on the availability of nutrients. Periodic fasting and other dietary restrictions can promote entry into a long‐lasting longevity program characterized by cellular protection and optimal function but can also activate regenerative processes that lead to rejuvenation, which are independent of the aging rate preceding the restricted period. Thus, a “juventology”‐based strategy can complement the traditional gerontology approach by focusing not on aging but on the longevity program affecting the life history period in which mortality is very low and organisms remain youthful, healthy, and fully functional.     

A Comprehensive Study of Soft Palate Development in Mice

Cleft palate is one of the most common congenital birth defects. Tremendous efforts have been made over the last decades towards understanding hard palate development. However, little is known about soft palate morphogenesis and myogenesis. Finding an appropriate surgical repair to restore physiological functions of the soft palate in patients with cleft palate is a major challenge for surgeons, and complete restoration is not always achievable. Here, we first analyzed the morphology, orientation and attachments of the four muscles of the murine soft palate and found that they are very similar to their counterparts in humans, validating the use of mus musculus as a model for future studies. Our data suggests that muscle differentiation extends from the lateral region to the midline following palatal fusion. We also detected an epithelial seam in the fusing soft palatal shelves, consistent with the process of fusion of the posterior palatal shelves, followed by degradation of the epithelial remnants. Innervation and vascularization are present mainly in the oral side of the soft palate, complementing the differentiated muscles. Cell lineage tracing using Wnt1-Cre;Zsgreen ^fl/fl mice indicated that all the tendons and mesenchyme embedding the soft palate muscles are neural crest-derived. We propose that the posterior attachment of the soft palate to the pharyngeal wall is an interface between the neural crest- and mesoderm-derived mesenchyme in the craniofacial region, and thus can serve as a potential model for the study of boundaries during development. Taken together, our study provides a comprehensive view of the development and morphology of the murine soft palate and serves as a reference for further molecular analyses.     

The distribution of syndecan during murine secondary palate morphogenesis.

The distribution of syndecan, an integral membrane proteoglycan, has been immunohistochemically mapped during the course of murine secondary palate morphogenesis, gestational days 12-15. Syndecan has been shown to mediate cell adhesion and shape change and to be involved in epithelial-mesenchymal interactions during the morphogenesis of several structures. Changes in epithelial cell architecture accompany and may serve to direct the reorientation of the murine secondary palatal shelves from a vertical position on either side of the tongue to a horizontal and adhering position above it. Using a monoclonal antibody made to the core protein of the ectodomain of syndecan, staining was observed to correlate with epithelial cell shape, packing and degree of differentiation. Staining of condensing mesenchyme was also observed. Syndecan may be involved in modulating epithelial cell shape, architecture and fates during both major phases of secondary palate morphogenesis: shelf reorientation and midline epithelial seam dissolution.     

Notch signal transduction: a real rip and more

The Notch signaling pathway functions in a wide variety of processes that regulate tissue patterning and morphogenesis in developing vertebrates and invertebrates. Research on the mechanism of ligand-induced Notch signal transduction has revealed a novel and essential element in the signal cascade. Some recent findings support a model in which sequential proteolytic cleavage serves to regulate Notch signal transduction.     

Bmpr1a signaling plays critical roles in palatal shelf growth and palatal bone formation

Cleft palate, including submucous cleft palate, is among the most common birth defects in humans. While overt cleft palate results from defects in growth or fusion of the developing palatal shelves, submucous cleft palate is characterized by defects in palatal bones. In this report, we show that the Bmpr1a gene, encoding a type I receptor for bone morphogenetic proteins (Bmp), is preferentially expressed in the primary palate and anterior secondary palate during palatal outgrowth. Following palatal fusion, Bmpr1a mRNA expression was upregulated in the condensed mesenchyme progenitors of palatal bone. Tissue-specific inactivation of Bmpr1a in the developing palatal mesenchyme in mice caused reduced cell proliferation in the primary and anterior secondary palate, resulting in partial cleft of the anterior palate at birth. Expression of Msx1 and Fgf10 was downregulated in the anterior palate mesenchyme and expression of Shh was downregulated in the anterior palatal epithelium in the Bmpr1a conditional mutant embryos, indicating that Bmp signaling regulates mesenchymal-epithelial interactions during palatal outgrowth. In addition, formation of the palatal processes of the maxilla was blocked while formation of the palatal processes of the palatine was significantly delayed, resulting in submucous cleft of the hard palate in the mutant mice. Our data indicate that Bmp signaling plays critical roles in the regulation of palatal mesenchyme condensation and osteoblast differentiation during palatal bone formation.     

Bicarbonate stimulated adenylyl cyclases

Bicarbonate ion is fundamental to the biology of all living organisms. HCO(3)(-) is vital to such diverse physiological processes as carbon fixation, cellular homeostasis, sperm maturation, and nucleotide synthesis. A defined subset of adenylyl cyclases identified in eukaryotes and prokaryotes are directly activated by HCO(3)(-). As such, cAMP represents the first identified biological effector for fluctuations in intracellular inorganic carbon levels. The identification of a signal transduction pathway activated by HCO(3)(-) has far reaching implications for understanding how the cell responds to fluctuations in this essential anion.     

How to Start a Wet Forest Ablaze: Perspectives on the Question of the Origins of Human Mindedness

This paper is a methodological and theoretical meditation on how some research has approached the question of the evolution of human cognitive traits. I discuss views that explicitly or implicitly endorse a view of human cognition as originating from a cause that can be singled out. Following Ross and Ladyman (2010), I suggest that this “singling-out” strategy correlates with a “container” metaphor that doesn’t fit with the interactive process-ontology of modern physics (Campbell 2009). Instead, Ross and Ladyman as well as Campbell recommend the metaphor of ‘emergence’. The logic and ontology of emergent systems finds resonance with developmental systems theory in biology. I suggest in agreement with Stotz (Phenomenology and the Cognitive Sciences 9(4), 2010) that we view the origins of human mindedness within such a framework.