Molecular insight into viral infection of Methanoarchaea and their respective viruses

该提案一般涉及甲烷古菌的病毒感染。第一部分和主要部分旨在从分子水平上研究和了解mazei Methanosarcina球面病毒（MetSV）与Methanosarcina球面病毒（Methanosarcina Spherical virus, MetSV）的相互作用。MetSV含有双链线性DNA，基因组大小为10,567bp，包含22个开放阅读框（orf），其中9个小orf编码长度小于50个氨基酸的蛋白质。为了在分子水平上深入了解病毒- M. mazei相互作用，我们通常会在提出的三个目标中应用生化和遗传方法的结合：(i)阐明感染周期内小蛋白的功能，特别是关注那些预测具有抗crispr或抗防御功能的蛋白。这将包括与Urska Repnik博士密切合作，通过透射电子显微镜（TEM）和扫描电子显微镜（SEM）研究和成像感染过程，并与Christel Kamp博士合作建立感染过程模型。（ii）阐明病毒DNA/RNA结合蛋白在感染过程中的预测调控功能。潜在的受调控宿主基因将通过对各自病毒突变体的RNAseq分析与野生型进行比较，以及与Julia Frunzke博士合作，通过染色质免疫沉淀测序（ChIP-SEq）定位宿主基因组的结合位点来鉴定。（三）通过计算目标预测工具和遗传方法确定病毒编码的srna并确定其特征，研究其在感染周期中的潜在功能。在第二部分中，我们的目标是开始研究存在于人类胃肠道中的甲烷古细菌病毒，以预测和未来评估甲烷古细菌病毒对人类肠道微生物群的影响。我们通过生物信息学方法在人类肠道微生物群宏基因组中鉴定出的绝大多数已鉴定但尚未表征的（methoano）古细菌病毒物种表明，古细菌病毒具有一种新的未被认识的多样性和重要性。因此，我们将与Cynthia M. Chibani博士和z项目合作，使用生物信息学方法表征已鉴定的新型病毒。同时，我们的目标是利用有针对性的方法从不同的样品中分离出已鉴定的甲烷古细菌病毒。从长远来看，在与Li Deng博士、michael Zimmermann博士和Bärbel Stecher-Letsch博士的合作下，这将使我们能够研究甲烷古细菌病毒对（人类）肠道微生物群的影响。

The proposal in general addresses viral infection of Methanoarchaea. The first and main part aims to study and understand the interaction between Methanosarcina mazei and its recent isolated lytic virus Methanosarcina Spherical Virus (MetSV) on a molecular level. MetSV contains double-stranded linear DNA with a genome size of 10,567bp containing 22 open reading frames (ORFs) including nine small ORFs encoding proteins with less than 50 amino acids in length. Aiming to gain insight into the virus - M. mazei interaction on the molecular level, we will generally apply a combination of biochemical and genetic approaches in the three goals proposed: (i) Elucidate the function of the small proteins within the infection cycle particularly focusing on those predicted to have anti-CRISPR or anti-defence function. This will include studying and imaging the infection process by transmission electron microscopy (TEM) and scanning electron microscopy (SEM) in close collaboration with Dr. Urska Repnik and modeling the infection process in collaboration with Dr. Christel Kamp. (ii) Illuminate the predicted regulatory function(s) of viral DNA/RNA binding proteins in the infection process. Potential regulated host genes will be identified by RNAseq analysis of respective viral mutants in comparison with the wild type, as well as by localizing binding sites of the host genome by chromatin immunoprecipitation sequencing (ChIP-SEq) in collaboration with Dr. Julia Frunzke. (iii) Identify and characterize the virus encoded sRNAs and study their potential functions during the infection cycle by computational target prediction tools and genetic approaches. In the second and minor part we aim to initiate studying methanoarchaeal viruses present in human gastrointestinal tracts to predict and in future also evaluate impacts of methanoarchaeal viruses on the human gut microbiome. The vast majority of identified yet uncharacterized (methoano)archaeal viral species we identified by bioinformatics approaches in metagenomes of human gut microbiomes suggests a novel unrecognized diversity and importance of archaeal viruses. Consequently, we will characterize the identified novel viruses using bioinformatics approaches in collaboration with Dr. Cynthia M. Chibani and the Z-project. In parallel we aim to isolate identified methanoarchaeal viruses from different samples using targeted approaches. In the long run and in collaboration with Dr. Li Deng, Dr. Micheal Zimmermann, and Dr. Bärbel Stecher-Letsch this will allow to study the impact of methanoarchaeal viruses on (human) gut microbiomes.