Abstract

The leaf extracellular space contains several peptidases, most of which are of unknown function. We isolated cDNAs for two extracellular serine carboxypeptidase III genes from tobacco (Nicotiana tabacum), NtSCP1 and NtSCP2, belonging to a phylogenetic clade not yet functionally characterized in plants. NtSCP1 and NtSCP2 are orthologs derived from the two ancestors of tobacco. Reverse transcription-polymerase chain reaction analysis showed that NtSCP1 and NtSCP2 are expressed in root, stem, leaf, and flower tissues. Expression analysis of the β-glucuronidase reporter gene fused to the NtSCP1 transcription promoter region confirmed this expression profile. Western blotting of NtSCP1 and expression of an NtSCP1-green fluorescent protein fusion protein showed that the protein is located in the extracellular space of tobacco leaves and culture cells. Purified His-tagged NtSCP1 had carboxypeptidase activity in vitro. Transgenic tobacco plants overexpressing NtSCP1 showed a reduced flower length due to a decrease in cell size. Etiolated seedlings of these transgenic plants had shorter hypocotyls. These data provide support for a role of an extracellular type III carboxypeptidase in the control of cell elongation.

Highlights

  • The leaf extracellular space contains several peptidases, most of which are of unknown function

  • The 5# and 3# ends were obtained by RACE-PCR, and the full coding region was amplified by reverse transcription (RT)-PCR using specific primers

  • Since tobacco is an amphitetraploid with two ancestors identified as Nicotiana sylvestris and Nicotiana tomentosiformis (Yukawa et al, 2006), we wondered whether NtSCP1 and NtSCP2 might be orthologs

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Summary

Introduction

The leaf extracellular space contains several peptidases, most of which are of unknown function. We isolated cDNAs for two extracellular serine carboxypeptidase III genes from tobacco (Nicotiana tabacum), NtSCP1 and NtSCP2, belonging to a phylogenetic clade not yet functionally characterized in plants. Etiolated seedlings of these transgenic plants had shorter hypocotyls These data provide support for a role of an extracellular type III carboxypeptidase in the control of cell elongation. The phylogenetic classification is supported by the structural data Both type I and II SCPs have two polypeptide chains formed from a single precursor polypeptide by the excision of a linker peptide of about 50 residues (Doan and Fincher, 1988), as in barley (Hordeum vulgare) carboxypeptidase I, which is cleaved into two polypeptide chains linked by disulfide bonds, forming the catalytically active heterodimer. Some SCPs are involved in the remobilization of nitrogen resources during seed germination, wound stress, and leaf senescence and may be involved in various signaling events important for plant growth and

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