Page 65 - Mouse Molecular Genetics

Full Abstracts
Program number is above title. Author in bold is the presenter.
in our C57BL/6J mouse, as expected. Modified Partek output was run through our newly developed Hotspot Detector for CNV
HD-CNV) software to identify 16 recurrent regions (CNV events overlapping each other by a minimum of 40 percent; between
samples). CNVs that were detected in only one tissue of a given individual, termed copy number changes, or CNCs, occurred in
both genotypes and tissues. The number of amplification CNCs was three times that of deletion CNCs. Six recurrent CNVs on
chromosome 8 overlapped with a previously observed CBA/CaJ CNV from the 351 dataset. This workflow, using two different
somatic tissues of the same mouse and novel software (HD-CNV) to detect and visualize recurrent CNVs, highlights patterns in
the dynamic murine genome that will give great insight into mechanisms underlying CNV formation and their phenotypic
Live cell imaging of random X-inactivation. Osamu Masui
Isabelle Bonnet
Patricia Le Baccon
Isabel Brito
Niall Murphy
Philippe Hupé
Emmanuel Barillot
Andrew Belmont
Haruhiko Koseki
Edith Heard
. 1)
Yokohama, Japan; 2) Curie Institute, Paris, France; 3) University of Illinois, Urbana, IL, USA.
Random X inactivation represents a paradigm for monoallelic gene regulation and epigenetic changes during early ES cell
differentiation. The choice of X chromosome to inactivate in XX cells is ensured by monoallelic regulation of Xist RNA via its
antisense transcription unit Tsix/Xite. Homologous pairing events have been proposed to underlie asymmetric Tsix expression
but this has never been addressed experimentally, owing to the dynamic and transient nature of such early developmental events.
Here we investigate the live cell dynamics and the outcome of Tsix pairing in differentiating mouse ES cells. We find an overall
increase in genome dynamics, including the Xics, during early ES cell differentiation. When they become paired however, Xic
loci show markedly reduced movements. Upon separation, Tsix expression becomes transiently monoallelic, thus providing a
window of opportunity for Xist up-regulation in cis to the silent Tsix allele. Our findings reveal the spatio-temporal choreography
of the two X chromosomes during early differentiation, and point to a direct role for pairing events in facilitating symmetry
breaking and monoallelic regulation of Xist during random X inactivation. Our recent live-cell imaging results will be also
presented and discussed.
Sphingosine-1-phosphate receptor 2 expression retains follicular helper T cells in germinal centers. Saya Moriyama
Noriko Takahashi
Jesse A. Green
Masato Kubo
Jason G. Cyster
Takaharu Okada
. 1)
Research Unit for
Immunodynamics, RCAI, RIKEN, Yokohama, Kanagawa, Japan; 2) Department of Microbiology and Immunology, University
of California, San Francisco, CA, USA; 3) Laboratory for Signal Network, RCAI, RIKEN, Yokohama, Kanagawa, Japan; 4)
Howard Hughes Medical Institute, University of California, San Francisco, CA, USA.
Follicular helper T (Tfh) cells are the helper T cell subset specialized in providing help to cognate antigen-specific B cells in
the secondary lymphoid organs, and thus play critical roles in humoral immune responses.Tfh cells are particularly important for
the germinal center (GC) reaction that is essential for long-term, high affinity antibody production. However, the molecular
mechanisms of how Tfh cells are physically associated with GCs are incompletely understood. Here we report that sphingosine-
phosphate receptor 2 (S1PR2), which has recently been shown to play a role in GC B cell localization, is also important for
GC-association of Tfh cells. Analysis using an S1pr2-reporter mouse strain suggested that S1PR2 is expressed at varied levels in
Tfh cells, and that S1PR2-high Tfh cells are localized in GCs whereas S1PR2-low Tfh cells are scattered throughout the B cell
follicle. S1PR2-deficient Tfh cells exhibited reduced accumulation in GCs, which was attributed to impaired retention in GCs
based on two-photon imaging analysis. Expression of Bcl6, Il4, and Il21 in Tfh cells was found to be positively correlated with
S1pr2 expression. These results suggest that S1PR2-high Tfh cells are GC-resident Tfh cells bearing the advanced capability to
promote long-term B cell responses.
A single-cell resolution Notch signaling reporter strain of mice. Sonja Nowotschin
Panos Xenopoulos, Evan Weiner, Kat
Hadjantonakis. Developmental Biology, Sloan-Kettering, New York, NY 10065, USA.
Live cell imaging is an essential tool for understanding the highly dynamic and coordinated events that drive cell lineage
specification and morphogenesis during mammalian development. To elucidate the critical role of signaling pathways and begin
to assay signal responsiveness, signaling reporter strains can be engineered by placing signaling responsive elements to direct the
expression of reporter genes. To date, several transgenic Notch reporter strains have been generated marking the sites of active
Notch signaling during development.
Here we report the construction of a fluorescent protein-based single-cell resolution Notch signaling reporter designed for live
visualization and tracking of individual cells
in vivo
in mouse embryos and adults and
ex vivo
in stem cells such as ES cells. We
have placed a CBF (also called RBP-Jk and CSL) responsive element (CBFRE) containing 4 copies of the mouse CBF1 binding
sites and an SV40 minimal promoter in front of a fluorescent protein fusion comprising human histone H2B linked to the Venus