Molecular mechanism of cadmium stress response in a traditional herbal medicine Anoectochilus roxburghii

Heavy metal toxicity is a serious constraint to orchids. Anoectochilus genus plants, important natural source for various active ingredients, are greatly sensitive to abiotic stress. To reveal the response mechanism of Anoec-tochilus genus to heavy metal stress, the transcriptomes of A. roxburghii seedlings under cadmium (Cd2+) treat-ment were analyzed. In total, seven heavy-metal-associated P1B-type ATPase (HMA) family genes were detected in A. roxburghii, including two Cd2+ treatment inducible HMA genes (ArHMA2 and ArHMA5.1). ArHMA2 and ArHMA5.1 contained classic 'E2-E2 ATPase' and 'Hydrolase' domains. Heterologous expression of ArHMA2 and ArHMA5.1 into the cadmium-sensitive yeast mutant strain (& UDelta;ycf1) confirmed that ArHMA2 and ArHMA5.1 might play roles in the tolerance of yeast to Cd2+ stress. To screen their regulators, promoter sequences of ArHMA2 and ArHMA5.1 were cloned, and several MYB and bHLH transcription factor (TF) recognition elements were iden-tified in their promoter regions. Furthermore, two Cd2+-induced TFs, ArMYB44.1 and ArbHLH28, were identi-fied. Both in vitro and in vivo assays confirmed that ArMYB44.1 and ArbHLH28 functioned in Cd2+ tolerance by up-regulating the expression of ArHMA2 and ArHMA5.1 genes. Our data revealed novel regulatory mechanism underlying the responses to Cd2+ stress and contributed to the breeding of orchids with greater environmental adaptability.

Automated summary

Heavy metal toxicity is a significant issue for orchids. This study investigated the response mechanism of Anoectochilus genus to heavy metal stress, and identified key genes and transcription factors involved in tolerance. These findings contribute to breeding orchids with better adaptability to adverse environments.