A data integration methodology for systems biology: Experimental verification

The integration of data from multiple global assays is essential to understanding dynamic spatiotemporal interactions within cells. In a companion paper, we reported a data integration methodology, designated Pointillist, that can handle multiple data types from technologies with different noise characteristics. Here we demonstrate its application to the integration of 18 data sets relating to galactose utilization in yeast. These data include global changes in mRNA and protein abundance, genome-wide protein–DNA interaction data, database information, and computational predictions of protein–DNA and protein–protein interactions. We divided the integration task to determine three network components: key system elements (genes and proteins), protein–protein interactions, and protein–DNA interactions. Results indicate that the reconstructed network efficiently focuses on and recapitulates the known biology of galactose utilization. It also provided new insights, some of which were verified experimentally. The methodology described here, addresses a critical need across all domains of molecular and cell biology, to effectively integrate large and disparate data sets

Proteomic identification of tmRNA substrates

The tmRNA–SmpB system releases ribosomes stalled on truncated mRNAs and tags the nascent polypeptides to target them for proteolysis. In many species, mutations that disrupt tmRNA activity cause defects in growth or development. In Caulobacter crescentus cells lacking tmRNA activity there is a delay in the initiation of DNA replication, which disrupts the cell cycle. To understand the molecular basis for this phenotype, 73 C. crescentus proteins were identified that are tagged by tmRNA under normal growth conditions. Among these substrates, proteins involved in DNA replication, recombination, and repair were overrepresented, suggesting that misregulation of these factors in the absence of tmRNA activity might be responsible for the delay in initiation of DNA replication. Analysis of the tagging sites within these substrates revealed a conserved nucleotide motif 5′ of the tagging site, which is required for wild-type tmRNA tagging