Enhancement of Ganoderic Acid Accumulation by Overexpression of an N-Terminally Truncated 3-Hydroxy-3-Methylglutaryl Coenzyme A Reductase Gene in the Basidiomycete Ganoderma lucidum

Abstract

Ganoderic acids produced by Ganoderma lucidum, a well-known traditional Chinese medicinal mushroom, exhibit antitumor and antimetastasis activities. Genetic modification of G. lucidum is difficult but critical for the enhancement of cellular accumulation of ganoderic acids. In this study, a homologous genetic transformation system for G. lucidum was developed for the first time using mutated sdhB, encoding the iron-sulfur protein subunit of succinate dehydrogenase, as a selection marker. The truncated G. lucidum gene encoding the catalytic domain of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) was overexpressed by using the Agrobacterium tumefaciens-mediated transformation system. The results showed that the mutated sdhB successfully conferred carboxin resistance upon transformation. Most of the integrated transfer DNA (T-DNA) appeared as a single copy in the genome. Moreover, deregulated constitutive overexpression of the HMGR gene led to a 2-fold increase in ganoderic acid content. It also increased the accumulation of intermediates (squalene and lanosterol) and the upregulation of downstream genes such as those of farnesyl pyrophosphate synthase, squalene synthase, and lanosterol synthase. This study demonstrates that transgenic basidiomycete G. lucidum is a promising system to achieve metabolic engineering of the ganoderic acid pathway.

A Novel Approach to Enhancing Ganoderic Acid Production by Ganoderma lucidum Using Apoptosis Induction

“…Methyl jasmonate and phenobarbital have also been demonstrated to increase GAs biosynthesis and expression of various biosynthetic genes [15], [16]. Over expression of HMGR in G. lucidum enhanced GA production indicating that HMGR play critical role for GA biosynthesis [39]. However, in this study, aspirin induced GAs production but reduced transcript of the LS and SQS.…”

Protoplasting, regeneration and transformation of medicinal mushroom Ganoderma multipileum using succinate dehydrogenase mutation gene as a selection marker

“…In fact, both these unique traits are significant obstacles impeding efficient transformation. These results contrasted greatly to several other studies on transformation in Basidiomycetes, i.e., Agaricus bisporus, Pleurotus ostreatus, G. lucidum, in which using the protoplast-polyethylene glycol (PEG) method, in combination with either GUS, hygromycin or bialaphos as an indicator or selection marker in normal dosage worked as expected (Case et al 1979;Sonnenberg et al 1988;Peng et al 1993;Li and Li 1999;Sun et al 2001;Kim et al 2004;Zhou et al 2008;Xu et al 2012).…”

Profiling and Quantifying Differential Gene Transcription Provide Insights into Ganoderic Acid Biosynthesis in Ganoderma lucidum in Response to Methyl Jasmonate

“…Despite significant research efforts to understand the influence of environmental factors on the GA biosynthesis [39], the regulatory networks by which environmental factors regulate GA biosynthesis remain unclear. Recent studies on genetically modified GA biosynthesis have focused on the genes restricted to the mevalonate pathway [21], [22]. Our previous study demonstrated that methyl jasmonate can significantly increase both the amount of GA and the transcription levels of known genes in the GA biosynthesis pathway [12].…”