ORIGINAL PAPER Heat shock modulates phosphorylation status and activity of nucleoside diphosphate kinase in cultured sugarcane cells Sunethra Dharmasiri • H. Michael Harrington • Nihal Dharmasiri Received: 1 July 2010 / Revised: 17 August 2010 / Accepted: 23 August 2010 / Published online: 7 September 2010 Ó Springer-Verlag 2010 Abstract Nucleoside diphosphate kinase (NDPK) is involved in the regeneration of nucleoside triphosphates (NTPs) through its phosphotransferase activity via an autophosphorylating histidine residue. Additionally, auto- phosphorylation of serine and/or threonine residues is documented for NDPKs from various organisms. However, the metabolic significance of serine/threonine phosphory- lation has not been well characterized. In this study we report the cloning and characterization of NDPKI from cultured sugarcane (Saccharum officinarum L. line H50- 7209) cells, and modulation of serine autophosphorylation of NDPK1 in response to heat-shock (HS). Heat-shock treatment at 40°C for 2 h resulted in a 40% reduction in labeled phosphoserine in NDPK1. This dephosphorylation was accompanied by an increase in NDPK enzyme activity. In contrast, NDPK1 in cultured tobacco (cv. W-38) cells did not show changes in autophosphorylation or increased enzyme activity in response to HS. The mRNA or protein level of NDPK1 did not increase in response to HS. Sug- arcane cells sustain the constitutive protein synthesis in addition to heat-shock protein synthesis during HS, while constitutive protein synthesis is significantly reduced in tobacco cells during HS. Thus, HS modulation of NDPK1 activity and serine dephosphorylation in sugarcane cells may represent an important physiological role in main- taining cellular metabolic functions during heat stress. Keywords Sugarcane Á Saccharum officinarum Á Heat-shock Á Nucleoside diphosphate kinase Á Serine phosphorylation Abbreviations NDP Nucleoside diphosphate HS Heat shock HSP Heat-shock protein HSR Heat shock response Introduction Nucleoside diphosphate kinase (abbreviated NDPK or NDK) (EC 2.7.4.6) is a ubiquitous enzyme that catalyzes the conversion of nucleoside diphosphates (NDPs) into nucleoside triphosphates (NTPs) through a ping-pong mechanism (Parks and Agarwal 1973). The reaction is mediated by a conserved histidine residue on the NDPK enzyme, and serves as the phosphorylated intermediate (Morera et al. 1995). In contrast to the early concepts of NDPK as a general housekeeping enzyme, later studies suggest physiologically essential multi-functional role for this enzyme (reviewed in Mehta and Orchard 2009). NDPKs associate with different sub-cellular structures, and form complexes with other proteins, as well as with nucleic acids (Thakur et al. 2009; Kavanaugh-Black et al. 1994; Engel et al. 1998; Galvis et al. 2001). The NDPK isoforms associated with chloroplasts, mitochondria, cytosol, plasma Communicated by Y. Lu. S. Dharmasiri (&) Á N. Dharmasiri Department of Biology, Texas State University, 601, University Drive, San Marcos, USA e-mail: sd20@txstate.edu H. M. Harrington Western Association of Agriculture Experiment Station Directors, Colorado State University, 16 Administration Building, Fort Collins, CO 80523, USA 123 Plant Cell Rep (2010) 29:1305–1314 DOI 10.1007/s00299-010-0917-6