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Molecular and Cellular Biology, June 2002, p. 4372-4382, Vol. 22, No. 12
0270-7306/02/$04.00+0     DOI: 10.1128/MCB.22.12.4372-4382.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Mitochondrial RNA Import in Leishmania tropica: Aptamers Homologous to Multiple tRNA Domains That Interact Cooperatively or Antagonistically at the Inner Membrane

Subhendra Nath Bhattacharyya, Saibal Chatterjee, and Samit Adhya*

Genetic Engineering Laboratory, Indian Institute of Chemical Biology, Calcutta 700032, India

Received 26 November 2001/ Returned for modification 15 January 2002/ Accepted 28 February 2002

A large number of cytoplasmic tRNAs are imported into the kinetoplast-mitochondrion of Leishmania by a receptor-mediated process. To identify the sequences recognized by import receptors, mitochondria were incubated with a combinatorial RNA library. Repeated cycles of amplification of the imported sequences (SELEX) resulted in rapid selection of several import aptamers containing sequence motifs present in the anticodon arm, the D arm, the V-T region, and acceptor stem of known tRNAs, confirming or suggesting the presence of import signals in these domains. As predicted, truncated derivatives of tRNAIle(UAU) containing the D arm or the V-T region were imported in vitro. Four aptamers were studied in detail. All were imported in vitro as well as in transiently transfected cells, using the same pathway as tRNA, but their individual import efficiencies were different. Two types of aptamers were discernible: the A arm and D arm homologues (type I), which were efficiently transferred across the inner mitochondrial membrane, and the V-T homologues (type II), which were not. Remarkably, subnanomolar concentrations of type I RNAs stimulated the entry of type II RNAs into the matrix, whereas type II RNAs inhibited inner membrane transfer of type I RNAs. Moreover, tRNATyr(GUA) and tRNAIle(UAU) interacted with one another as type I and type II, respectively. Such cooperative and antagonistic interactions may allow the use of a limited number of receptors to recognize a large number of tRNAs of variable affinity and enable the maintenance of a properly balanced tRNA pool for mitochondrial translation.

* Corresponding author. Mailing address: Genetic Engineering Laboratory, Indian Institute of Chemical Biology, 4 Raja S. C. Mullick Rd., Calcutta 700032, India. Phone: 91 33 473 3491, ext. 136. Fax: 91 33 473 5197/0284. E-mail: sadhya{at}iicb.res.in.

Molecular and Cellular Biology, June 2002, p. 4372-4382, Vol. 22, No. 12
0022-538X/02/$04.00+0     DOI: 10.1128/MCB.22.12.4372-4382.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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