Pax3 encodes a paired homeobox containing transcription factor that is expressed in neuroepithelium, neural crest, and presomitic mesoderm (1). Homozygous mouse embryos carrying a loss-of-function Pax3 allele (Pax3 Ϫ/Ϫ ) develop open neural tube defects, such as exencephaly or spina bifida (2), and die around embryonic day 14 (E14) 4 as a consequence of heart defects (3). More recently, Pax3 has been shown to function at the nodal point in melanocyte stem cell differentiation (4). Heterozygous embryos (Pax3 ϩ/Ϫ ) are viable but exhibit white patches on their bellies caused by defective development of neural crest-derived melanocytes. This suggests that disruption of the Pax3-dependent developmental program may cause defects in the development of neural crest-derived structures.Two main Pax3-binding sites have been identified and are found in most Pax3 target elements as follows: (i) a binding site derived from the Drosophila Paired (ATTA N5 GTTCC), and (ii) a Pax3 paired domain binding site (CGTCAC(G/A)(C/ G)TT) identified by Epstein et al. (5) by CASTing (DNA-binding site selection assays) in the c-Met promoter region. In addition several paired domains, such as GTTCC, CAGTGT, GTTAT, GTGTGA, and CAAGG (6), as well as the homeodomain ATTA, have been suggested to be "putative Pax3-binding motifs" (7,8). More recently, Corey and Underhill (9) demonstrated that Pax3 can regulate target genes through alternative modes of DNA recognition. They observed that although the microphthalmia-associated transcription factor element is characterized by suboptimal recognition motifs for the paired domain and homeodomain, it sustains a higher level of Pax3 binding than TRP-1, which contains a canonical paired domain site. The basis for this difference involved a context-dependent cooperative binding event requiring both the paired and homeodomain, whereas the paired domain alone was sufficient for TRP-1 recognition.Since Pax3 is important in diverse cellular functions during development, we wanted to identify additional genes regulated by Pax3. To accomplish this we utilized oligonucleotide arrays and RNA isolated from Pax3-transfected cell lines and promoter-based data mining (6). Based on the putative Pax3-binding

A DBA/2J (D2) transgenic mouse line with cyan fluorescent protein (CFP) reporter expression in ganglion cells was developed for the analysis of ganglion cells during progressive glaucoma. The Thy1-CFP D2 (CFP-D2) line was created by congenically breeding the D2 line, which develops pigmentary glaucoma, and the Thy1-CFP line, which expresses CFP in ganglion cells. Microsatellite marker analysis of CFP-D2 progeny verified the genetic inclusion of the D2 isa and ipd loci. Specific mutations within these loci lead to dysfunctional melanosomal proteins and glaucomatous phenotype in D2 mice. Polymerase chain reaction analysis confirmed the inclusion of the Thy1-CFP transgene. CFP-fluorescent ganglion cells, 6–20 μm in diameter, were distributed in all retinal regions, CFP processes were throughout the inner plexiform layer, and CFP-fluorescent axons were in the fiber layer and optic nerve head. Immunohistochemistry with antibodies to ganglion cell markers NF-L, NeuN, Brn3a, and SMI32 was used to confirm CFP expression in ganglion cells. Immunohistochemistry with antibodies to amacrine cell markers HPC-1 and ChAT was used to confirm weak CFP expression in cholinergic amacrine cells. CFP-D2 mice developed a glaucomatous phenotype, including iris disease, ganglion cell loss, attrition of the fiber layer, and elevated intraocular pressure. A CFP-D2 transgenic line with CFP-expressing ganglion cells was developed, which has (1) a predominantly D2 genetic background, (2) CFP-expressing ganglion cells, and (3) age-related progressive glaucoma. This line will be of value for experimental studies investigating ganglion cells and their axons in vivo and in vitro during the progressive development of glaucoma.

Cryptococcal meningitis remains a significant opportunistic infection in HIV-infected individuals worldwide, despite availability of antiretroviral therapies in developed nations. Current therapy with amphotericin B is difficult to administer and only partially effective. Mechanisms of cryptococcal neuropathogenesis are still not clearly defined. In the present study, we used a C57Bl/6 mouse model with intravenous inoculation of three isogenic strains of Cryptococcus neoformans: H99, Cap59, and Pkr1-33. These strains differ in their capsule production and are normocapsular, hypocapsular, and hypercapsular, respectively. We studied the role of capsule in the morbidity and mortality of our host animal. Surprisingly, we found that the hypercapsular strain was least virulent while the strains that produced less capsule were more virulent and had higher concentrations of organism in the brain. These results suggest that neurovirulence is related to total capsule volume and rate of capsule accumulation in the brain, rather than the amount of capsule produced per organism. Therapies which decrease central nervous system dissemination and inhibit replication rates in the brain may be more effective than therapies which target capsule production.

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