Worm Breeder's Gazette 14(1): 74 (October 1, 1995)
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
|1||Division of Molecular Biology, The Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX Amsterdam, The Netherlands.|
|2||National Cancer Institute, Frederick Cancer Res. Dev. Fac., Frederick, MD 21702, USA.|
Upon exposure to natural toxins tumor cells can become resistant to structurally and functionally unrelated drugs (multidrug resistance or MDR). The classical form of MDR is caused by a P-glycoprotein (P-gp) encoded by the human MDR1 gene. Recently it has been found that MDR can also be conferred by the Multidrug resistance associated protein (MRP), a member of the ABC family of transporters. In C. elegans there is besides the P-gp gene family also an MRP gene family. We investigate whether the proteins encoded by these mrp genes are involved in the protection of the nematode against natural occurring compounds. The MRP gene has been first identified in a drug resistant small cell lung cancer cell line. The protein is located in the plasma-membrane and confers resistance by decreasing the intracellular drug concentration or the intracellular distribution. The predicted protein consists of a N-terminal half of eight transmembrane domains and an ATP-binding site and a C-terminal half of four transmembrane domains and an ATP binding site. The MRP mRNA can be detected in all human tissues. The intracellular distribution differs from that of the P- glycoproteins; P-glycoproteins are predominantly expressed in the apical membranes of epithelial cells whereas MRP is mainly found in intra-cellular vesicles. Recent studies suggest that also the mechanism of action of MRP differs from that of P-gp. Mrp can also function as a transporter of glutathione conjugated substrates. It is thought that MRP is involved in (acquired) resistance against drugs beyond the classic MDR spectrum of drugs. The human MRP is most related to the Leishmania tarentolae P-glycoprotein ltpgpA and the Yeast CF1 gene. Both can confer resistance to a number of heavy metal ions. So far there are no clues for the natural physiological function of the MRP protein. In C. elegans four mrp gene family members have been identified; mrp-1, -2, -3 and -4. mrp-1 was first identified as a EST clone from Ventner and Waterston's sequencing effort. The other three genes were identified as cDNA clones from Y. Kohara's sequencing project. We started to analyze mrp-1 and mrp-2. Both genes are localized on the X-chromosome next to each other, the nucleotide sequence determination has almost been completed. The predicted protein sequence of both genes resembles that of the human MRP; they are both 71% similar at the amino acid level. Using RT-PCR we found that mrp-1 in expressed in all developmental stages. The highest expression was found in early larval stages. To investigate the gene's function we isolated a Tc1 allele and a deletion allele of mrp-1. In the mrp-1 deletion mutant 3 kb genomic DNA has been deleted containing several transmembrane domains and the first ATP-binding site. The mrp-1 knock-out was found to be sensitive to heavy metals like cadmium and arsenite then wild-type animals (see fig 1).