Regulatory Mechanisms in Stem Cell Biology

Stem cells are a subject of intense and increasing interest because of their biological properties and potential medical importance. Unfortunately, the field has been difficult for the nonspecialist to penetrate, in part because of ambiguity about what exactly constitutes a stem cell. A working definition is useful in order to pose the important questions in stem cell biology. However, since different people define stem cells in different ways (for examples, see37 and 76), formulating a generally acceptable definition can lead to a conclusion similar to that of U. S. Supreme Court Justice Byron White's in regard to pornography: “It's hard to define, but I know it when I see it.” A minimalist definition is that stem cells have the capacity both to self-renew and to generate differentiated progeny. Although this is in many respects inadequate, it immediately highlights some important problems: How at each cell division is a stem cell able to pass on its “stem” properties to at least one of its two daughters? And what determines whether stem cell divisions will be self-renewing, or differentiating

Generation of Induced Pluripotent Stem Cells from the Prairie Vole

The vast majority of animals mate more or less promiscuously. A few mammals, including humans, utilize more restrained mating strategies that entail a longer term affiliation with a single mating partner. Such pair bonding mating strategies have been resistant to genetic analysis because of a lack of suitable model organisms. Prairie voles are small mouse-like rodents that form enduring pair bonds in the wild as well as in the laboratory, and consequently they have been used widely to study social bonding behavior. The lack of targeted genetic approaches in this species however has restricted the study of the molecular and neural circuit basis of pair bonds. As a first step in rendering the prairie vole amenable to reverse genetics, we have generated induced pluripotent stem cell (IPSC) lines from prairie vole fibroblasts using retroviral transduction of reprogramming factors. These IPSC lines display the cellular and molecular hallmarks of IPSC cells from other organisms, including mice and humans. Moreover, the prairie vole IPSC lines have pluripotent differentiation potential since they can give rise to all three germ layers in tissue culture and in vivo. These IPSC lines can now be used to develop conditions that facilitate homologous recombination and eventually the generation of prairie voles bearing targeted genetic modifications to study the molecular and neural basis of pair bond formation

Integration of multiple instructive cues by neural crest stem cells reveals cell-intrinsic biases in relative growth factor responsiveness

Growth factors can influence lineage determination of neural crest stem cells (NCSCs) in an instructive manner, in vitro. Because NCSCs are likely exposed to multiple signals in vivo, these findings raise the question of how stem cells would integrate such combined influences. Bone morphogenetic protein 2 (BMP2) promotes neuronal differentiation and glial growth factor 2 (GGF2) promotes glial differentiation; if NCSCs are exposed to saturating concentrations of both factors, BMP2 appears dominant. By contrast, if the cells are exposed to saturating concentrations of both BMP2 and transforming growth factor β1 (which promotes smooth muscle differentiation), the two factors appear codominant. Sequential addition experiments indicate that NCSCs require 48–96 hrs in GGF2 before they commit to a glial fate, whereas the cells commit to a smooth muscle fate within 24 hr in transforming growth factor β1. The delayed response to GGF2 does not reflect a lack of functional receptors; however, because the growth factor induces rapid mitogen-activated protein kinase phosphorylation in naive cells. Furthermore, GGF2 can attenuate induction of the neurogenic transcription factor mammalian achaete-scute homolog 1, by low doses of BMP2. This short-term antineurogenic influence of GGF2 is not sufficient for glial lineage commitment, however. These data imply that NCSCs exhibit cell-intrinsic biases in the timing and relative dosage sensitivity of their responses to instructive factors that influence the outcome of lineage decisions in the presence of multiple factors. The relative delay in glial lineage commitment, moreover, apparently reflects successive short-term and longer-term actions of GGF2. Such a delay may help to explain why glia normally differentiate after neurons, in vivo

Odor-evoked activity in the MOE and VNO is required for female sexual receptivity.

<p>(A) No difference between single or double mutant and control females in the length of the estrus cycle. (B) Males showed comparable frequency of anogenital chemoinvestigation (sniffing) to females of all genotypes. (C) A majority of females of all genotypes were mounted by WT males in ≥50% of assays. (D) Chemosensory control of female sexual receptivity. There was a significant effect on receptivity from both <i>Cnga2</i> and <i>Trpc2</i>, with no interaction between the two, resulting in a significant decrease in sexual receptivity in <i>Cnga2^-/-</i>, <i>Trpc2^-/-</i> double mutant females. Two-way ANOVA: <i>Cnga2 F</i>(1,56)  = 4.05, p = 0.049; <i>Trpc2 F</i>(1,56)  = 6.03, p = 0.017; interaction <i>F</i>(1,56)  = 0.077, p = 0.782. Mean ± SEM; *p<0.05; N≥8/cohort.</p

Odor-evoked activity in the MOE is not required for male-typical sexual displays.

<p>(A) <i>Trpc2^-/-</i> and <i>Cnga2^-/-</i>, <i>Trpc2^-/-</i> females were equally likely to mount a male and significantly different from control females who never mounted males. (B) <i>Cnga2^-/-</i>, <i>Trpc2^-/-</i> females mount males earlier than <i>Trpc2^-/-</i> females. t-test <i>t</i>(13)  = −2.543, p = 0.025. (C) <i>Cnga2^-/-</i>, <i>Trpc2^-/-</i> females mount males more often than <i>Trpc2^-/-</i> females. t-test <i>t</i>(13)  = 3.28, p = 0.006. Mean ± SEM. *p<0.05. N≥11/cohort.</p

Complex Chemosensory Control of Female Reproductive Behaviors

<div><p>Olfaction exerts a profound influence on reproductive physiology and behavior in many animals, including rodents. Odors are recognized by sensory neurons residing in the main olfactory epithelium (MOE) and the vomeronasal organ (VNO) in mice and many other vertebrates. The relative contributions of the MOE and VNO in the display of female behaviors are not well understood. Mice null for <i>Cnga2</i> or <i>Trpc2</i> essentially lack odor-evoked activity in the MOE and VNO, respectively. Using females mutant for one or both of <i>Cnga2</i> and <i>Trpc2</i>, we find that maternal care is differentially regulated by the MOE and VNO: retrieval of wandering pups requires the MOE and is regulated redundantly by the VNO whereas maternal aggression requires both sensory epithelia to be functional. Female sexual receptivity appears to be regulated by both the MOE and VNO. <i>Trpc2</i> null females have previously been shown to display male-type mounting towards other males. Remarkably, we find that females double mutant for <i>Cnga2</i> and <i>Trpc2</i> continue to mount other males, indicating that the disinhibition of male-type sexual displays observed in <i>Trpc2</i> null females does not require chemosensory input from a functional MOE. Taken together, our findings reveal a previously unappreciated complexity in the chemosensory control of reproductive behaviors in the female mouse.</p></div

Alternative Neural Crest Cell Fates Are Instructively Promoted by TGFβ Superfamily Members

How growth factors influence the fate of multipotent progenitor cells is not well understood. Most hematopoietic growth factors act selectively as survival factors, rather than instructively as lineage determination signals. In the neural crest, neuregulin instructively promotes gliogenesis, but how alternative fates are determined is unclear. We demonstrate that bone morphogenic protein 2 (BMP2) induces the basic–helix-loop-helix protein MASH1 and neurogenesis in neural crest stem cells. In vivo, MASH1^+ cells are located near sites of BMP2 mRNA expression. Some smooth muscle differentiation is also observed in BMP2. A related factor, transforming growth factor β1 (TGFβ1), exclusively promotes smooth muscle differentiation. Like neuregulin, BMP2 and TGFβ1 act instructively rather than selectively. The neural crest and hematopoietic systems may therefore utilize growth factors in different ways to generate cellular diversity

Neural control of sexually dimorphic behaviors

All sexually reproducing animals exhibit gender differences in behavior. Such sexual dimorphisms in behavior are most obvious in stereotyped displays that enhance reproductive success such as mating, aggression, and parental care. Sexually dimorphic behaviors are a consequence of a sexually differentiated nervous system, and recent studies in fruit flies and mice reveal novel insights into the neural mechanisms that control these behaviors. In the main, these include a diverse array of novel sex differences in the nervous system, surprisingly modular control of various stereotyped dimorphic behavioral routines, and unanticipated sensory and central modulation of mating. We start with a brief overview to provide the appropriate conceptual framework so that the advances made by the newer studies discussed subsequently can be fully appreciated. We restrict our review to reporting progress in understanding the basis of mating and aggression in fruit flies and mice

Chapter Eight Turning ON Caspases with Genetics and Small Molecules

Caspases, aspartate-specific cysteine proteases, have fate-determining roles in many cellular processes including apoptosis, differentiation, neuronal remodeling, and inflammation (for review, see Yuan &amp; Kroemer, 2010). There are a dozen caspases in humans alone, yet their individual contributions toward these phenotypes are not well understood. Thus, there has been considerable interest in activating individual caspases or using their activity to drive these processes in cells and animals. We envision that such experimental control of caspase activity can not only afford novel insights into fundamental biological problems but may also enable new models for disease and suggest possible routes to therapeutic intervention. In particular, localized, genetic, and small-molecule-controlled caspase activation has the potential to target the desired cell type in a tissue. Suppression of caspase activation is one of the hallmarks of cancer and thus there has been significant enthusiasm for generating selective small-molecule activators that could bypass upstream mutational events that prevent apoptosis. Here, we provide a practical guide that investigators have devised, using genetics or small molecules, to activate specific caspases in cells or animals. Additionally, we show genetically controlled activation of an executioner caspase to target the function of a defined group of neurons in the adult mammalian brain