Although only 44% identical to human karyopherin a,, human karyopherin a2 (Rchl protein) substituted for human karyopherin a, ( (3, 4). Indeed, mapping of the docking site(s) for Nup98 showed it (them) to be located in the repeat-containing N-terminal half of Nup98 (12). Finally, the repeat-containing nucleoporins also contain binding sites for plO (M. Matunis, G.B., and M.H., unpublished data). It has been proposed that the repeatcontaining nucleoporins serve as a stationary phase and the transport factors as the mobile phase in transport across the NPC (12).We showed previously that the a subunit serves as an NLS receptor and proposed that the X3 subunit serves as an adaptor that binds to the a subunit-NLS substrate complex and to the repeat-containing nucleoporins (4). Here we show that the previously used karyopherin a subunit [corresponding to hSRP-1/NPI-1 (14, 15)], now termed karyopherin a1, can be replaced by karyopherin a2 [corresponding to the hSRP-1-related Rchl (16)]. Although karyopherin a, and a2 are only 44% identical, we did not detect any functional difference. We also show that karyopherin 3 is localized in the cytoplasm and at the nuclear rim and is largely lost after digitonin permeabilization of cells. Unlike karyopherin a, which alone cannot bind to the nuclear envelope of digitonin-permeabilized cells, karyopherin , can bind without karyopherin a being present. In an import reaction with all recombinant transport factors, the karyopherin a subunits, Ran, and plO are shown to enter the nucleus, whereas the ,B subunit remains at the nuclear envelope. This result suggests that the heterodimeric karyopherin complex is dissociated during transport. Overlay binding assays showed that the ,B subunit binds to the repeatcontaining nucleoporins. These data are consistent with the previously proposed function of karyopherin J3 as an adaptor between karyopherin a/NLS substrate complex and repeatcontaining nucleoporins.Abbreviations: HSA, human serum albumin; NLS, nuclear localization sequence; NPC, nuclear pore complex; FITC, fluorescein isothiocyanate.

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We have isolated five cDNA clones for rat liver catalase (hydrogen peroxide:hydrogen peroxide oxidoreductase, EC 1.11.1.6). These clones overlapped with each other and covered the entire length of the mRNA, which had been estimated to be 2.4 kilobases long by blot hybridization analysis of electrophoretically fractionated RNA. Nucleotide sequencing was carried out on these five clones and the composite nucleotide sequence of catalase cDNA was determined. The 5' noncoding region contained 83 bases and was followed by 1581 bases of an open reading frame that encoded 527 amino acids. The 3' noncoding region was 831 bases long and contained long repeats of the unit AC. The amino acid sequence deduced from the nucleotide sequence of the cDNAs showed about 90% homology with the reported primary structure of bovine liver catalase. The molecular weight of rat liver catalase was calculated to be 59,758 from the predicted amino acid sequence. The amino acid residues in contact with the heme group are completely identical for bovine liver and rat liver catalases. The amino acid sequence at the COOH terminus was confirmed by the results ofcarboxypeptidase P treatment of the protein purified from rat liver in the presence of leupeptin. Rat liver catalase has no cleavable signal peptide for translocation of the enzyme into peroxisomes.

Since hepatitis C virus (HCV), a major causative agent of posttransfusional non-A, non-B hepatitis, is a positive stranded RNA virus, it is supposed to replicate via a negative RNA strand. Although strand specific reverse transcription-polymerase chain reaction (RT-PCR) method was recently developed to detect each strand of HCV RNA, the specificity of the strategy has remained to be determined. In this study, using in vitro transcribed positive and negative stranded HCV RNAs mixed with hepatic cellular RNA from normal liver, we found that this strategy did not distinguish between the two RNA strands, but that chemical modification of RNA samples at the 3' end followed by strand specific RT-PCR made specific detection possible. Liver tissues, sera and peripheral blood mononuclear cells (PBMC) from ten patients with chronic HCV infection were analyzed with the novel strategy of RT-PCR combined with RNA modification. Positive and negative strands of HCV RNA were detected in liver tissues of ten (100%) and nine (90%) cases, respectively. Negative RNA strand was detected also in sera of five cases (50%), positive strand being detected in nine cases (90%). In PBMC, positive strand of HCV RNA was detected in eight cases (80%), whereas negative strand in only one case (10%), suggesting that HCV has much less cellular tropism to PBMC than to hepatocytes.