The physiological role of prion protein (PrP) remains unknown. Mice devoid of PrP develop normally but are resistant to scrapie; introduction of a PrP transgene restores susceptibility to the disease. To identify the regions of PrP necessary for this activity, we prepared PrP knockout mice expressing PrPs with amino-proximal deletions. Surprisingly, PrP lacking residues 32-121 or 32-134, but not with shorter deletions, caused severe ataxia and neuronal death limited to the granular layer of the cerebellum as early as 1-3 months after birth. The defect was completely abolished by introducing one copy of a wild-type PrP gene. We speculate that these truncated PrPs may be nonfunctional and compete with some other molecule with a PrP-like function for a common ligand.

Mice devoid of PrP are resistant to scrapie and fail to replicate the agent. Introduction of transgenes expressing PrP into such mice restores susceptibility to scrapie. We find that truncated PrP devoid of the five copper binding octarepeats still sustains scrapie infection; however, incubation times are longer and prion titers and protease-resistant PrP are about 30-fold lower than in wild-type mice. Surprisingly, brains of terminally ill animals show no histopathology typical for scrapie. However, in the spinal cord, infectivity, gliosis, and motor neuron loss are as in scrapie-infected wild-type controls. Thus, while the region comprising the octarepeats is not essential for mediating pathogenesis and prion replication, it modulates the extent of these events and of disease presentation.

Expression of the transmembrane glycoprotein CD98 (encoded by SLC3A2) is increased in intestinal inflammatory conditions, such as inflammatory bowel disease (IBD), and in various carcinomas, yet its pathogenetic role remains unknown. By generating gain-and loss-of-function mouse models with genetically manipulated CD98 expression specifically in intestinal epithelial cells (IECs), we explored the role of CD98 in intestinal homeostasis, inflammation, and colitis-associated tumorigenesis. IEC-specific CD98 overexpression induced gut homeostatic defects and increased inflammatory responses to DSS-induced colitis, promoting colitis-associated tumorigenesis in mice. Further analysis indicated that the ability of IEC-specific CD98 overexpression to induce tumorigenesis was linked to its capacity to induce barrier dysfunction and to stimulate cell proliferation and production of proinflammatory mediators. To validate these results, we constructed mice carrying conditional floxed Slc3a2 alleles and crossed them with Villin-Cre mice such that CD98 was downregulated only in IECs. These mice exhibited attenuated inflammatory responses and resistance to both DSS-induced colitis and colitis-associated tumorigenesis. Together, our data show that intestinal CD98 expression has a crucial role in controlling homeostatic and innate immune responses in the gut. Modulation of CD98 expression in IECs therefore represents a promising therapeutic strategy for the treatment and prevention of inflammatory intestinal diseases, such as IBD and colitis-associated cancer.

SummaryA new congenital haemorrhagic diathesis is described. It is characterized by the poor and slaw wound healing, profuse and long bleeding and by probable absence of spontaneous and joint bleeding. It is due to the deficiency of a factor which makes the fibrin clots insoluble in urea. This factor is probably the fibrin stabilizing factor of Laki and Lorand, although no proof of their identity can yet be given. The deficiency appears in both sexes and is probably an autosomal recessive disease.

Summary: Conventional approaches to produce transgenic mice recurrently yield unpredictable patterns and levels of transgene expression, a situation calling for the development of new techniques to overcome these drawbacks in the context of overexpression studies. Here we present an efficient method for rapid and reproducible transgenesis using the recombinase mediated cassette exchange (RMCE) (Bouhassira et al.: Blood 90:3332-3344, 1997) procedure. A lox511-EGFP-TK/neoloxP cassette was placed under the control of the endogenous mouse b-actin promoter. Heterozygous mice revealed strong and ubiquitous EGFP expression throughout embryogenesis and adulthood. Reproducibly, the same expression pattern was obtained with RMCE when it was used to replace the EGFP-harboring cassette by ECFP or placental alkaline phosphatase (PLAP) reporter genes (DePrimo et al.: Transgenic Res 5:459-466, 1996). Furthermore, the RMCE procedure proved efficient as well in embryonic stem (ES) Key words: AGRP; targeted transgenesis; RMCE; mouse; lox511As a consequence of unpredictable positional effects and varying copy numbers, transgenic mice, generated by conventional DNA injection techniques, commonly present incoherent transgene expression patterns. Therefore, it is often necessary to analyze multiple mouse lines before conclusions, as to the effect of overexpressed transgenic products, can be drawn. In contrast, targeting transgenes into defined and precharacterized loci yields predictable expression patterns due to the invariable transcriptional control exerted by the given endogenous regulatory sequences.In order to reproducibly obtain strong and ubiquitous transgene expression in mice, we have therefore focused investigations on endogenous b-actin, a locus consistently expressed during embryonic development and adulthood. The b-actin genomic region was cloned by screening a BAC library with intron 3 and used to generate the targeting vector by inserting a lox511-EGFPTKNeo-loxP cassette at the b-actin translation start codon in exon 2 (Fig. 1a). Strong EGFP expression was observed in the targeted BALB/c ES-cell clones, as well as in mice which were generated from these ( Fig. 2). Mice heterozygous for the targeted b-actin-EGFP allele were fertile and presented strong, ubiquitous EGFP fluorescence throughout embryogenesis (starting at embryonic day (E)2.5) (Fig. 2b) and in adult organs such as brain, eye, heart, liver, lung, kidney, muscle, thymus, spleen, small intestine, stomach, large intestine, skin, uterus, and tail (Fig. 2a). Except for a 10% lower body weight, X-ray analysis, blood chemo-gram, hemogram, urine analysis, and histopathology all failed to reveal differences with respect to wildtype animals. Homozygous mice, however, were not viable and died between days E10.5 and E11.5 (Fig. 2b) (and as published previously;Shawlot et al., 1998).The b-actin-EGFP ES cells were used to test the efficiency of Cre-recombinase-mediated cassette exchange (RMCE) of lox511-loxP flanked transgenes.Heterospecific lox-sites (wildtype loxP...

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