Main Goal

SPP 2002 – Small proteins in Prokaryotes, an unexplored world

Prokaryotes are highly abundant and diverse and live in literally all ecological niches. They have broad impact on the environment and our health, and are crucial for biotechnology and the food industry. To fully understand their versatile lifestyles and exploit their metabolic capacities, we must know their genetic / biochemical repertoires and cellular regulatory processes. Modern genomics and transcriptomics have over the past decade discovered a wealth of hidden small genes containing short open reading frames (sORFs) in many prokaryotic genomes. These sORFs encoded small proteins of <50 amino acids in length, and are typically missed by automated gene predictions. Preliminary studies have shown that these new small proteins impact disparate cellular processes, running the gamut of energy generation, transport, virulence, symbiosis, sporulation, and photosynthesis. They often localize to membranes and can modulate the activity of larger protein complexes. These initial findings notwithstanding, the full repertoire and function of this cellular small proteome comprising perhaps hundreds of small proteins in any given prokaryote remains to be uncovered.

The small proteins have been difficult to detect due to technical limitations but new technologies are emerging to enable their global profiling and systematic investigation of their physiological roles. For example, deep-sequencing based translatome analyses (ribosome profiling) can now tell us how many potential sORFs are translated in an organism. Likewise, new proteomics / peptidomics techniques can reveal novel small proteins and quantitate their expression. Our nationwide priority program aims to unravel this emerging major class of prokaryotic gene products. We will focus on the prokaryotic small proteome due to the unique membranes and dynamics of protein complexes in bacteria and archaea, as well as their environmental impact. Besides, they are ideally suited as model organisms attributably to their simple cellular structure devoid of compartmentation, availability of excellent tools for fast genetic manipulation and phenotypic analysis.

We aim to 1) globally identify small proteins in selected prokaryotes using comparative genomics, translatomics and peptidomics, 2) advance novel analytical tools to identify small proteins and their physiological roles, and 3) elucidate the functions and interaction partners of small proteins as well as underlying molecular mechanisms. Exploring the small proteome in Prokaryotes will not only deepen our understanding of gene expression regulation and protein function in general, but may also provide new innovative biotechnological applications and antimicrobial strategies. By disclosing representative functions and mechanisms for the products of this major class of genes in prokaryotic genomes we envisage to uncover novel unique prokaryotic principles in regulation of gene expression and assembly/­disassembly of large (membrane) complexes.