Universal GFP reporter for the study of vascular development.

We report the generation and characterization of transgenic mouse and zebrafish expressing green fluorescent protein (GFP) specifically in vascular endothelial cells in a relatively uniform fashion. These reporter lines exhibit fluorescent vessels in developing embryos and throughout adulthood, allowing visualization of the general vascular patterns with single cell resolution. Furthermore, we show the ability to purify endothelial cells from whole embryos and adult organs by a single step fluorescence activated cell sorting. We expect that these transgenic reporters will be useful tools for imaging vascular morphogenesis, global gene expression profile analysis of endothelial cells, and high throughput screening for vascular mutations.

A C. elegans Ror receptor tyrosine kinase regulates cell motility and asymmetric cell division.

Ror kinases are a family of orphan receptors with tyrosine kinase activity that are related to muscle specific kinase (MuSK), a receptor tyrosine kinase that assembles acetylcholine receptors at the neuromuscular junction. Although the functions of Ror kinases are unknown, similarities between Ror and MuSK kinases have led to speculation that Ror kinases regulate synaptic development. Here we show that the Caenorhabditis elegans gene cam-1 encodes a member of the Ror kinase family that guides migrating cells and orients the polarity of asymmetric cell divisions and axon outgrowth. We find that tyrosine kinase activity is required for some of the functions of CAM-1, but not for its role in cell migration. CAM-1 is expressed in cells that require its function, and acts cell autonomously in migrating neurons. Overexpression and loss of cam-1 function result in reciprocal cell-migration phenotypes, indicating that levels of CAM-1 influence the final positions of migrating cells. Our results raise the possibility that Ror kinases regulate cell motility and asymmetric cell division in organisms as diverse as nematodes and mammals.

Identification of Caenorhabditis elegans genes required for neuronal differentiation and migration.

To understand the mechanisms that guide migrating cells, we have been studying the embryonic migrations of the C. elegans canal-associated neurons (CANs). Here, we describe two screens used to identify genes involved in CAN migration. First, we screened for mutants that died as clear larvae (Clr) or had withered tails (Wit), phenotypes displayed by animals lacking normal CAN function. Second, we screened directly for mutants with missing or misplaced CANs. We isolated and characterized 30 mutants that defined 14 genes necessary for CAN migration. We found that one of the genes, ceh-10, specifies CAN fate. ceh-10 had been defined molecularly as encoding a homeodomain protein expressed in the CANs. Mutations that reduce ceh-10 function result in Wit animals with CANs that are partially defective in their migrations. Mutations that eliminate ceh-10 function result in Clr animals with CANs that fail to migrate or express CEH-23, a CAN differentiation marker. Null mutants also fail to express CEH-10, suggesting that CEH-10 regulates its own expression. Finally, we found that ceh-10 is necessary for the differentiation of AIY and RMED, two additional cells that express CEH-10.