The structure and regulation of homeologous tobacco endochitinase genes of Nicotiana sylvestris and N. tomentosiformis origin.
The fungicidal class I chitinases are believed to be important in the induced defense response of plants. We isolated and partially characterized genomic clones representing two members, CHN14 and CHN50, of the gene subfamily encoding these enzymes in Nicotiana tabacum L. cv. Havana 425. The coding sequences of genes CHN14, CHN50, and CHN48, which was cloned earlier, are identical at 79-95% of the positions. Tobacco is an amphidiploid species derived from ancestors most closely related to the present-day species N. sylvestris and N. tomentosiformis. Southern analysis of genomic DNA, comparison of deduced amino acid sequences, and partial sequencing of the purified enzymes suggest that the gene pairs CHN48/CHN50 and CHN14/CHN14' are homeologues. Gene CHN48, which encodes chitinase A (Mr ca. 34 kDa), and gene CHN14 are derived from N. tomentosiformis; whereas gene CHN50, which encodes chitinase B (Mr ca. 32 kDa), and gene CHN14' are derived from N. sylvestris. Class I chitinases are induced in leaves of plants treated with ethylene or infected with the fungal pathogen Cercospora nicotianae and in cultured cells transferred to medium without added auxin and cytokinin. RNase protection assays show that under these conditions transcripts encoded by the homeologues CHN48 and CHN50 account for greater than 90% of the total chitinase mRNA. The less abundant transcript, CHN48, consistently showed a greater degree of induction than CHN50. Expression of the homeologues CHN14 and CHN14' represented less than 10% of the total chitinase mRNA. They showed a pattern of hormonal regulation similar to CHN48 and CHN50, but transcripts of these genes were not detected in leaves infected with C. nicotianae. Therefore the two sets of homeologues are regulated in the same way by hormones and respond differently to infection by a pathogen.
van Buuren
M
M
Friedrich Miescher-Institut, Basel, Switzerland.
Neuhaus
J M
JM
Shinshi
H
H
Ryals
J
J
Meins
F
F
D000595:Amino Acid Sequence; D015139:Blotting, Southern; D002688:Chitinases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D005786:Gene Expression Regulation; D017343:Genes, Plant; D015698:Genomic Library; D008969:Molecular Sequence Data; D005810:Multigene Family; D010947:Plants, Toxic; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D012689:Sequence Homology, Nucleic Acid; D013045:Species Specificity; D014026:Nicotiana / Q000201:enzymology / Q000235:genetics*
D012333:RNA, Messenger; D011994:Recombinant Proteins; D004247:DNA; D002688:Chitinases