No abstract

Summary Multidrug resistance (MDR) in cancer cells is a major contributor to the failure of chemotherapy treatment. This paper describes a novel protein named the anthracycline resistance associated (ARA) protein.The ara gene is amplified in the MDR leukaemia line CCRF-CEM/E1000 and its mRNA is overexpressed. ARA belongs to the ATP binding cassette (ABC) family of proteins. Another ABC protein, the multidrug resistance-associated protein (MRP), has previously been reported to be overexpressed in the CEM/E1000 subline. The primary amino acid sequence of ARA indicates that it is 49.5 kDa without glycosylation, and that it has one potential glycosylation site. ARA has one ATP binding site and associated transmembrane regions. This is in contrast to MRP (190 kDa, 172 kDa deglycosylated) and most other higher eukaryote ABC proteins, which consist of two similar halves, each having one ATP binding site. In addition to ARA being coexpressed with MRP, comparison of amino acid sequences showed that, among known proteins, ARA is most similar to the C-terminal half of MRP.

In each of 5 tomato (Lycopersicon esculentum Mill) cultivars examined, alcohol dehydrogenase (E.C.1 .I .I .I) specijic activity decreased during the early stages of ripening and then increased in the postclimacteric period. Alcohol dehydrogenase spec$c activity also increased when immature, mature or pink fruit were placed in an atmosphere of 3% (VIV) oxygen. Measurements of oxygen in the internal tissues and the respiratory quotient throughout ripening established that fruit ripening in air does not suffer an oxygen stress. Increases in alcohol dehydrogenase activity were also observed in pink fruit exposed to 10% (VIV) carbon dioxide. Such atmospheres are known to result in a lowering of the cytoplasmic pH in plants and it is suggested that alcohol dehydrogenase activity is induced during ripening in response to cytoplasmic acid stress. The increased ADH activity during ripening may contribute to flavor development.

A cDNA library was constructed from RNA from the pericarp of ripe tomato fruit and four cDNAs encoding ADH2 were isolated and characterized. The cDNAs encode a peptide 379 amino acids in length. They hybridized strongly with a 1.8 kb RNA species well represented in RNA from ripe, but not from mature, unripe fruit, and strongly to a similar RNA species present in hypoxic, but not in aerobic roots. Northern analysis showed that the mRNA for ADH2 in fruit increased in abundance through ripening, particularly during late ripening. In pericarp tissue of fruit, the Adh2 mRNA level increased to a maximum within 8-16 h of exposure to atmospheres with 3 ~o (v/v) oxygen, and returned to the basal level within 16 h of a return to air. The mRNA level was sensitive to the oxygen level in the atmosphere, increasing 20-fold in 12~o (v/v) oxygen and 100-fold in 3~o oxygen.The homologous tomato Adh2 gene was isolated from a genomic library. The gene has an overall length of 2334 bp from transcription start site to poly(A) addition site and includes eight introns.Southern blot analysis of tomato genomic DNA identified multiple Adh2-related sequences. Two of these, PSA1 and PSA2, were cloned and found to have 94~o similarity with each other and 77~o similarity with the tomato Adh2 gene over a 1000 bp region. The homologous regions include introns and exons but the equivalent exons contain frame shifts, deletions and stop codons. The two regions are therefore presumptive pseudogenes.Abbreviations: ADH, alcohol dehydrogenase; Adh, pertaining to an ADH gene; ARE, anaerobic responsive element.