Atsushi Yuki

Activation of Glutathione Transferase P Gene by Lead Requires Glutathione Transferase P Enhancer I

Glutathione transferase P (GST-P) is specifically induced in rat liver and kidney by lead cation. The increase of GST-P mRNA after lead administration is blocked by actinomycin D, suggesting that GST-P production by lead is regulated at the transcriptional level. To further determine which part of the flanking region of the GST-P gene has the lead-responsive cis-element in vivo, we utilized transgenic rats with five different constructs having GST-P and/or chloramphenicol acetyltransferase coding sequence. We studied the effect of lead on these transgenic rats and on transfected NRK (normal rat kidney) cells and found that GST-P induction by lead is indeed regulated at the transcriptional level and that the GST-P enhancer I (GPEI) enhancer is an essential cis-element required for the activation of the GST-P gene by lead. GPEI consists of two AP-1 (c-Jun/c-Fos heterodimer) site-like sequences that are palindromically arranged and can bind AP-1. c-jun mRNA in the liver increased after lead administration and GST-P, and c-Jun had patchy expression in the same hepatocytes 24 h after lead exposure. These results suggest that activation of the GST-P gene by lead is mediated in major part by enhancer GPEI and that AP-1 may be involved at least partially. GPEI has been shown to have essential sequence information for the trans-activation of the GST-P gene during chemical hepatocarcinogenesis of the rat (Morimura, S., Suzuki, T., Hochi, S., Yuki, A., Nomura, K., Kitagawa, T., Nagatsu, I., Imagawa, M., and Muramatsu, M.(1993) Proc. Natl. Acad. Sci. U. S. A. 90, 2065-2068; Suzuki, T., Imagawa, M., Hirabayashi, M., Yuki, A., Hisatake, K., Nomura, K., Kitagawa, T., and Muramatsu, M. (1995) Cancer Res. 55, 2651-2655). The present study establishes that the same enhancer element does operate in the activation of the GST-P gene by lead regardless of the trans-activators involved.

Successful production of transgenic rats

Abstract DNA of pSV2‐gpt‐gE1A or SV2‐cat microinjected into pronuclei of fertilized rat eggs was found incorporated into chromosomes of 3 out of 48 (6.3%) or 10 out of 66 (15.2%) new born rats, respectively. The transgenic rats carrying SV2‐cat DNA transmitted the transgenes to their G1 progeny.

Fate of exogenous DNA carried into mouse eggs by spermatozoa.

Abstract Mouse spermatozoa carried pSV2‐gpt DNA in the culture medium Into eggs at fertilization in vitro. The exogenous DNA in the fertlized eggs could be detected by PCR analysis even at their hatched biastocyst stages. However, none of 130 mice resulted from such embryos was transgenic, while microinjection of the DNA into pronuclei of fertilized mouse eggs produced 13 transgenic mice out of 146 born.