Completion of the DNA sequences of the human genome and that of the nematode Caenorhabditis elegans allows the large-scale identification and analysis of orthologs of human genes in an organism amenable to detailed genetic and molecular analyses. We are determining gene expression profiles in specific cells, tissues, and developmental stages in C. elegans. Our ultimate goal is not only to describe detailed gene expression profiles, but also to gain a greater understanding of the organization of gene regulatory networks and to determine how they control cell function during development and differentiation. The use of C. elegans as a platform to investigate the details of gene regulatory networks has several major advantages. Two key advantages are that it is the simplest multicellular organism for which there is a complete sequence (C. elegans Sequencing Consortium 1998), and it is the only multicellular organism for which there is a completely documented cell lineage (Sulston and Horvitz 1977; Sulston et al. 1983). C. elegans is amenable to both forward and reverse genetics (for review, see Riddle et al. 1997). A 2-week life span and generation time of just 3 days for C. elegans allows experimental procedures to be much shorter, more flexible, and more cost-effective compared to the use of mouse or zebrafish models for genomic analyses. Finally, the small size, transparency, and limited cell number of the worm make it possible to observe many complex cellular and developmental processes that cannot easily be observed in more complex organisms. Morphogenesis of organs and tissues can be observed at the level of a single cell (White et al. 1986). As events have shown, investigating the details of C. elegans biology can lead to fundamental observations about human health and biology (Sulston 1976; Hedgecock et al. 1983; Ellis and Horvitz 1986). We are using complementary approaches to examine gene expression in C. elegans. We are constructing transgenic animals containing promoter green fluorescent protein (GFP) fusions of nematode orthologs of human genes. These transgenic animals are examined to determine the time and tissue expression pattern of the promoter::GFP constructs. Concurrently, we are undertaking serial analysis of gene expression (SAGE) on all developmental stages of intact animals and on selected purified cells. Tissues and selected cells are isolated using a fluorescence activated cell sorter (FACS) to sort promoter::GFP marked cell populations. To date we have purified to near homogeneity cell populations for embryonic muscle, gut, and a subset of neurons. The SAGE and promoter::GFP expression data are publicly available at http://elegans.bcgsc.bc.ca.

Astrocyte activation is a hallmark of central nervous system injuries resulting in glial scar formation (astrogliosis). The activation of astrocytes involves metabolic and morphological changes with complex underlying mechanisms, which should be defined to provide targets for astrogliosis intervention. Astrogliosis is usually accompanied by an upregulation of glial fibrillary acidic protein (GFAP). Using an in vitro scratch injury model, we scratched primary cultures of cerebral cortical astrocytes and observed an influx of calcium in the form of waves spreading away from the wound through gap junctions. Using the calcium blocker BAPTA-AM and the JNK inhibitor SP600125, we demonstrated that the calcium wave triggered the activation of JNK, which then phosphorylated the transcription factor c-Jun to facilitate the binding of AP-1 to the GFAP gene promoter to switch on GFAP upregulation. Blocking calcium mobilization with BAPTA-AM in an in vivo stab wound model reduced GFAP expression and glial scar formation, showing that the calcium signal, and the subsequent regulation of downstream signaling molecules, plays an essential role in brain injury response. Our findings demonstrated that traumatic scratch injury to astrocytes triggered a calcium influx from the extracellular compartment and activated the JNK/c-Jun/AP-1 pathway to switch on GFAP expression, identifying a previously unreported signaling cascade that is important in astrogliosis and the physiological response following brain injury.

In vertebrates, Fibroblast Growth Factors (FGFs) and their receptors are involved in various developmental and pathological processes, including neoplasia. The number of FGFs and their large range of activities have made the understanding of their precise functions di cult. Investigating their biology in other species might be enlightening. A sequence encoding a putative protein presenting 30 ± 40% identity with the conserved core of vertebrate FGFs has been identi®ed by the C. elegans sequencing consortium. We show here that this gene is transcribed and encodes a putative protein of 425 amino acids (aa). The gene is expressed at all stages of development beyond late embryogenesis, peaking at the larval stages. Loss-of-function mutants of the let-756 gene are rescued by the wild type fgf gene in germline transformation experiments. Two partial loss-of-function alleles, s2613 and s2809, have a mutation that replaces aa 317 by a stop. The truncated protein retains the FGF core but lacks a C-terminus portion. These worms are small and develop slowly into clear and scrawny, yet viable and fertile adults. A third allele, s2887, is inactivated by an inversion that disrupts the ®rst exon. It causes a developmental arrest early in the larval stages. Thus, in contrast to the other nematode fgf gene egl-17, let-756/fgf is essential for worm development.

Water movement plays vital roles in both physiological and pathological conditions in the brain. Astrocytes are responsible for regulating this water movement and are the major contributors to brain edema in pathological conditions. Aquaporins (AQPs) in astrocytes play critical roles in the regulation of water movement in the brain. AQP1, 3, 4, 5, 8, and 9 have been reported in the brain. Compared with AQP1, 4, and 9, AQP3, 5, and 8 are less studied. Among the lesser known AQPs, AQP5, which has multiple functions identified outside the central nervous system, is also indicated to be involved in hypoxia injury in astrocytes. In our study, AQP5 expression could be detected both in primary cultures of astrocytes and neurons, and AQP5 expression in astrocytes was confirmed in 1- to 4-week old primary cultures of astrocytes. AQP5 was localized on the cytoplasmic membrane and in the cytoplasm of astrocytes. AQP5 expression was downregulated during ischemia treatment and upregulated after scratch-wound injury, which was also confirmed in a middle cerebral artery occlusion model and a stab-wound injury model in vivo. The AQP5 increased after scratch injury was polarized to the migrating processes and cytoplasmic membrane of astrocytes in the leading edge of the scratch-wound, and AQP5 over-expression facilitated astrocyte process elongation after scratch injury. Taken together, these results indicate that AQP5 might be an important water channel in astrocytes that is differentially expressed during various brain injuries.