Tools for genetic engineering of the yeast Hansenula polymorpha

Hansenula polymorpha is a methylotrophic yeast species that has favorable properties for heterologous protein production and metabolic engineering. It provides an attractive expression platform with the capability to secrete high levels of commercially important proteins. Over the past few years many efforts have led to advances in the development of this microbial host including the generation of expression vectors containing strong constitutive or inducible promoters and a large array of dominant and auxotrophic markers. Moreover, highly efficient transformation procedures used to generate genetically stable strains are now available. Here, we describe these tools as well as the methods for genetic engineering of H. polymorpha

A global characterisation of the archaeal transcription machinery

Archaea employ a eukaryote-like transcription apparatus to transcribe a bacteria-like genome; while the RNA polymerase, basal factors and promoter elements mirror the eukaryotic RNA polymerase II system, archaeal genomes are densely packed with genes organised into multicistronic transcription units. The molecular mechanisms of archaeal transcription have been studied and characterised in great detail in vitro, but until recently relatively little was known about its global characteristics. In this chapter we discuss an integrated view of transcription from the molecular to the global level. Systems biology approaches have provided compelling insights into promoter and terminator DNA elements, the genome-wide distribution of transcription initiation- and elongation factors and RNA polymerase, the archaeal transcriptome and chromatin organisation. Overall these analyses illuminate transcription from a genome-wide perspective and serve as a resource for the community. In addition, Big Data can often validate mechanistic models based on biochemical and structural information, and generate new working hypotheses that can be thoroughly tested and dissected in vitro. This is an exciting time to study gene expression in the archaea since we are at the brink of a comprehensive yet detailed understanding of transcription