We have been studying the effect of Zn and Cu on binding of Nuclear Hormone Receptors (NHRs)to DNA and effect on thyroid hormone regulated transcription. Our results show that both Zn and Cu have positive effect on both DNA and transcription regulation at concentrations around 10-5 M but affect negatively the transcription at higher concentrations. We have also observed direct binding of 67Cu to DBD of jellyfish RXR. Our results are keeping with the possibility that DBD of NHRs may be ble to quire a second conformation under the effect of divalent ions with d orbitals. This may reflect a physiological role of these ions in modulation of function of NHRs as well as the possibility of an alternative conformation which may be involved in mechanism of NHR function such as their release from DNA during the receptor recycling. Keeping with the possibility of physiological role of divalent metals in regulation of development, the effect of Cu was reversed by natural chelator of Cu, histidine. We have been studying the function of an enzyme active in histidine production, carnosinase in worms. We have found that carnosinase inhibition results in developmental arrest. We currently are studying the function of methionine, which is conserved in DNA binding domain of NHRs and we propose that this methionine may be conserved for its side chain flexibility termed methionine flipping, which is likely to be essential for NHR function. We have been also working on mechanism of action of CHR3, a nuclear hormone receptor in C. elegans, which is involved in regulation of molting. We have shown that CHR3 is a critical regulator of all four molts in C. elegans development. In search for genes downstream of CHR3, we have obtained a line expressing dpy-7::gfp and found that the fluorescence significantly decreased after introducing CHR3 RNAi by feeding. We have also identified that the homologue of SKIP is a cofactor in CHR3 regulation and recently proved that this cofactor is involved in multiple regulatory pathways in C. elegans development. We used a new method of mapping C. elegans mutations by PCR so called snip-SNP mapping to determine mapping of molt-4 mutation. With this method we confirmed that the mutant molt-4 is mapped to chromosome III. We continue our work on a cubmomedusan jellyfish T. cystphora. We have continual laboratory culture of T. cystophora larvae and specialized structures containing eyes - the rhopalia and we are searching for NHRs. We started a small scale of cDNA sequencing project. We have also identified second NHR in T. cystophora and we are currently trying to clone it. This receptor is apparently derived from RXR/RAR/TR subfamily of NHRs. T. cystophora, a highly developed jellyfish with eyes possessing single layer of cells which are photoreceptors as well as pigment cells seems to be important for understanding evolution of Metazoa.
| Kostrouchova, Marta; Housa, Daniel; Kostrouch, Zdenek et al. (2002) SKIP is an indispensable factor for Caenorhabditis elegans development. Proc Natl Acad Sci U S A 99:9254-9 |
| Kostrouchova, M; Krause, M; Kostrouch, Z et al. (2001) Nuclear hormone receptor CHR3 is a critical regulator of all four larval molts of the nematode Caenorhabditis elegans. Proc Natl Acad Sci U S A 98:7360-5 |
