In order to identify novel target molecules for the treatment of neovascular eye diseases, we performed comprehensive transcriptome analyses in postnatal mouse retinas, because a considerable number, if not all, of endothelial genes in developing retinal vessels are also expected to be expressed in pathological angiogenesis. To date, a series of high-throughput analyses exploiting cultured or tissue-derived ECs have shown that endothelial gene expression fluctuates depending on microenvironments 
. This notion prompted us to isolate retinal ECs from Tie2GFP
Tg mice, which can reduce alterations in gene expression by omitting time-consuming antibody labeling steps. Indeed, the bioinformatics analysis of our microarray data using the NIH-DAVID software demonstrated that most of the gene ontology terms (level 3 biological processes with p
) enriched in the 1,623 endothelial genes were associated with dynamic cell behavior such as “cell motility”, “cell migration”, and “angiogenesis” (Table S2
). However, we should be aware of drawbacks inherent in our FACS and microarray analyses. Firstly, the expression levels of short-life genes, such as those up-regulated under hypoxia, may be affected during the sample preparation. Secondly, endothelial genes expressed at limited sites, such as sprouting vascular tips 
, might be masked by the lack of expression in the majority of ECs. Thirdly, the relative comparison of gene expression levels between ECs and non-ECs does not necessarily extract EC-specific genes. To overcome these obstacles, the exact expression pattern of particular genes should be determined by highly sensitive procedures, such as the use of reporter mouse strains.
While 80 RhoGEFs comprising the Dbl family and the Dock family have been identified 
, we successfully discovered that Arhgef15 acts as an EC-specific GEF to activate Cdc42 downstream of VEGF signaling. Although endothelial Arhgef15 expression was also detected in embryonic vasculature (Figure S3B
), previous literature reported Arhgef15 expression in vSMCs 
and in neurons 
. Thus, the Arhgef15 expression should further be examined in various tissues during development and in adults. In addition to Cdc42 activation, Arhgef15 potentiated RhoJ inactivation, which may underlie VEGF-induced RhoJ inactivation 
. In this process, a distinct GTPase activating protein 
downstream of Arhgef15 or Cdc42, may directly inactivate RhoJ. The full understanding of the Arhgef15-mediated signaling cascades will provide further information for the discovery of novel drug targets.
Based on the impaired vascular growth in Arhgef15
-KO retinas, we propose that Arhgef15 is a potential molecular target for selectively manipulating angiogenesis without affecting tissue homeostasis. Given the lower collapse rate of HUVECs induced by Arhgef15 knockdown than by Cdc42 knockdown, it seems likely that multiple GEFs, such as Fgd5 
, are involved in Cdc42 activation in ECs. It is also possible that Arhgef15 modulates the activity of alternative Rho small GTPases, such as RhoA 
. Therefore, enhanced anti-angiogenic effects may be achieved by targeting multiple endothelial G proteins. For this purpose, our transcriptome profiling is a useful source of information to identify novel target molecules for the treatment of neovascular eye diseases, as well as diseases such as cancer characterized by deregulated angiogenesis.