基于微生物共培养的隐性基因簇激活策略

Co-culture strategy for activating the cryptic gene cluster in microorganisms

摘要:

微生物次级代谢产物是药物先导化合物的重要源泉之一。随着测序技术的迅猛发展，越来越多的微生物基因组得以测序完成。伴随着测序技术的进步，生物信息学也得到了快速发展。基因组序列分析发现，链霉菌和丝状真菌等微生物中存在大量的已知的或未知的次级代谢物生物合成基因簇(secondary metabolite-biosynthetic gene clusters，SM-BGCs)。然而，在实验室培养条件下大部分基因簇无法表达或表达量很低，导致难以发现这些基因簇所对应的代谢产物，人们将这类基因簇称为“隐性基因簇”或“沉默基因簇”。通过调节基因簇中特异调控基因或基因簇外全局性调控基因的表达，对代谢途径的定向改造，以及将基因簇导入异源宿主等策略，能够激活部分隐性基因簇的表达。通过激活隐性基因簇的表达，能够发现通过常规实验室培养无法获得的具有独特生物活性的新结构代谢产物，成为创新药物的重要来源之一。然而，这些基因簇激活策略都严重依赖于对特定菌株或宿主的遗传操作。近年来，通过模拟自然混合培养中微生物间相互作用，开发了通过混合特定微生物菌株在厌氧或好氧条件下激活隐性基因簇的方法，称之为共培养激活策略。这种策略不依赖于基因组信息，也不依赖于对特定菌株或宿主的遗传操作技术，具有操作简单和方便的优点。共培养策略需要混合培养的不同微生物具有相似的生长速度以及不能够产生拮抗等要求，因而也部分限制了该策略的应用。近期合成微生物组学的出现有可能改变这一限制，使共培养策略得到更加广泛的应用。本文围绕微生物共培养体系和应用、基于共培养策略的产物挖掘以及可能的激活机制等进行了综述。

Abstract:

Microbial secondary metabolites are the rich sources of lead compounds. With the booming of genome sequencing techniques, increasing microbial genomes have been sequenced and the corresponding bioinformatics has also developed rapidly. Based on the bioinformatics analysis, a large number of secondary metabolite-biosynthetic gene clusters (SM-BGCs) have been discovered in filamentous microorganisms such as Streptomyces and filamentous fungi. However, most of SM-BGCs are dormant or silent under the conventional culture conditions with their corresponding metabolites difficult to be detected, which are regarded as cryptic or silent gene cluster. Through manipulation of the specific regulatory genes in the cluster or global regulatory genes outside the cluster, reconstruction of the metabolic pathways and heterologous expression in other species can activate the expression of cryptic gene clusters. Through activating expression of the cryptic gene clusters, researchers can discover new structural metabolites with unique bioactivities that cannot be obtained through conventional laboratory culture. Activating the cryptic gene clusters is a key approach to produce lead compounds. However, such activation strategies are heavily dependent on the genetic manipulation of the specific strains. Recently, researchers employ co-culture of specific microbial strains under anaerobic or aerobic conditions to activate the cryptic SM-BGCs by mimicking the microbial interactions occurring in natural environments. This strategy does not rely on the genomic information or the genetic manipulation of target strains, and it has the advantage of easy operation. The co-culture strategy requires that the different microorganisms in the mixed culture have similar growth rates and no antagonistic interaction, which partially limits its application. Emergence of the synthetic microbiomes may overcome this limitation and make the co-culture strategy more widely used in future. Here, we overviewed the co-culture systems and applications, the mining of natural products specifically produced in microbial co-culture, and the possible mechanisms underlying this phenomenon.