MAP kinase phosphorylation is dispensable for cell division, but required for cell growth in Drosophila.

Proper activation of the Ras/MAPK pathway is broadly required during development, and in many cases, signal transduction downstream of the receptor is linear. Thus, different mechanisms exist to properly regulate the large number of specific developmental outputs that are required by the activation of this pathway. Previously, we have reported a regulated cytoplasmic sequestration of phosphorylated MAPK (pMAPK) in developing Drosophila compound eyes and wings "called MAPK Cytoplasmic Hold". In the developing wing, we have shown that cytoplasmic hold promotes the differentiation of wing vein tissue, while pMAPK nuclear translocation regulates growth and division. We had also suggested that the Ras pathway signals for inducing cell growth and cell division split upstream of the nuclear translocation of MAPK itself. Here, we further refine the role of MAPK in Drosophila. We report evidence that suggests, for the first time, that the phosphorylation of MAPK is itself another step in the regulation of cell growth and division in both Drosophila wing and eye cells. We show that inhibition of MAPK phosphorylation, or pMAPK nuclear translocation, is sufficient to block cell growth, but not cell division. These data suggest that non-phosphorylated MAPK is sufficient to induce cell division, but not cell growth, once inside the nucleus of the cell.

Characterization of gene expression profiles of normal canine retina and brain using a retinal cDNA microarray.

PURPOSE: Construction of a canine retinal custom cDNA microarray for comprehensive retinal gene expression profiling and application for the identification of genes that are preferentially expressed in the retina and brain lobes using a brain pool reference tissue. METHODS: A cDNA microarray was constructed utilizing clones obtained from a normalized canine retinal expressed sequence tag library. Gene expression profiles were analyzed for normal retina, as well as the cortex of the frontal, occipital, and temporal brain regions. Each sample was studied against a reference sample of pooled brain RNA. Data from a quantified scanned image were normalized using the loess subgrid procedure. Retina-enriched genes were identified using the Significance Analysis of Microarrays (SAM) algorithm, and confirmed by northern blot analyses for selected genes. Differences between biological samples were displayed using principal component analysis (PCA). RESULTS: Expression profiles for each tissue set were analyzed against the common reference of pooled brain. Changes in expression between the sample and the reference were higher in the retina (27.9%) than the individual brain tissues (2-6.6%). Furthermore, all individual retinal samples were clearly separated from any of the hybridizations using brain tissue in the PCA. The accuracy of observed changes in expression has been confirmed by northern blot analysis using five randomly chosen genes that represented a wide range of different expression levels between retina and brain. CONCLUSIONS: We have established an accurate and robust microarray system suitable for the investigation of expression patterns in the retina and brain. Characterization of the gene expression profiles in normal retina will facilitate the understanding of the processes that underline differences between normal and diseased retinas.