Characterization of a basic chitinase which is secreted by cultured pumpkin cells

Abstract

chemical causing callus formation, suggesting that it may be A basic chitinase was secreted into culture medium of pumpkin cell suspension cultures. The chitinase was purified from independent of the presence of 2,4-D. Perhaps, induction is caused by osmotic or wounding stress. Levels of chitinase the culture medium. A cDNA encoding the pumpkin chitinase mRNA markedly increased at 4 h after transfer of pumpkin was cloned by reverse transcription (RT)-PCR and rapid amplification of cDNA ends (RACE) methods. The chitinase callus cells into fresh culture liquid medium. They were also high at later stages of cell suspension culture. In transgenic gene was strongly expressed in pumpkin callus cells, but little or not at all in mature leaf, young leaf, cotyledon, stem, tobacco BY-2 cells, into which the pumpkin chitinase cDNA hypocotyl and root of pumpkin. No chitinase mRNA was was introduced, the recombinant pumpkin chitinase was expressed and secreted into the culture medium, suggesting that detected in intact pumpkin fruit tissues. However, chitinase was induced during callus formation from sliced pumpkin fruit the signal peptide of pumpkin chitinase also functions for secretion from tobacco BY-2 cells. tissues. Induction also occurred in the absence of 2,4-D, a perhaps by releasing endogenous factors that are related to nodulation factors (De Jong et al. 1992, Vijn et al. 1993.In plants, four classes of chitinases have so far been proposed on the basis of their primary structures (Collinge et al. 1993). Class I chitinases contain an amino-terminal cysteine-rich domain of about 40 amino acids, a chitinbinding domain that is homologous to hevein and wheat germ agglutinin, and a highly conserved main structure, separated by a variable hinge region. Most class I chitinases have a basic isoelectric point and are localized in vacuoles. Indeed, many prochitinases have vacuolar targeting signals, i.e seven carboxy-terminal extra amino acids (Neuhaus et al. 1991). Class II chitinases are similar to class I chitinases but lack an amino-terminal cysteine-rich domain. They appear to be acidic proteins and are located in the apoplast. Class III chitinases have sequences very different from most of class I and class II chitinases. They can be acidic or basic proteins, and may be localized in the apoplast. Class IV chitinases contain a cysteine-rich domain and a conserved main structure, resembling class I chitinases, but they are significantly smaller as a consequence of four major deletions.

Chitinase production in pine callus ( Pinus sylvestris L.): a defense reaction against endophytes?

“…Specific chitinases belonging to class II are produced during somatic embryogenesis (De Jong et al 1992;Mo et al 1996;Dong and Dunstan 1997;Domon et al 2000), and chitinases are also found in cell-suspension cultures (Kragh et al 1991;Masuta et al 1991;Arie et al 2000).…”

Molecular cloning and expression of drought-induced protein 3 (DIP3) encoding a class III chitinase in upland rice

“…Chitinases are divided into seven classes (I-VII) according to their structure, substrate specificity, mechanism of catalysis, and sensitivity to inhibitors (Arie et al, 2000). Chitinases have some domains based on their amino acid sequences.…”

Purification, characterization of a CkChn134 protein from Cynanchum komarovii seeds and synergistic effect with CkTLP against Verticillium dahliae

“…Callus and suspension culture was expressed both with and without 2,4-D presence in the nutrient medium. 22 In D. japonica callus culture, exogenous ethylene elevated the chitinase activity. 23 In addition, jasmonic acid (JA) acts as signal that promotes defense system in plant and regulates the plant's responses to wounding.…”

“…25 Furthermore, wounding and osmotic stress induce the synthesis of basic extracellular chitinase in Cucuribia sp. 22 In high salt, low temperature, and drought treatment, the transcripts of CkChn134 were upregulated [ Fig. 7(C-E)].…”