Limits to a classic paradigm: Most transcription factors regulate genes in multiple biological processes

Transcription factors (TFs) are important drivers of cellular decision-making. When bacteria encounter a change in the environment, transcription factors alter the expression of a defined set of genes in order to adequately respond. It is commonly assumed that genes regulated by the same TF should be involved in the same biological process. Examples of this are methods that rely on coregulation to infer function of not yet annotated genes. We have previously shown that only 21% of TFs regulate functionally homogeneous genes based on the proximity of their catalyzed reactions in the metabolic network. Here, we provide more evidence to support the claim that a one TF/one process relationship is not a general property. We show that the observed functional heterogeneity of regulons is not a result of the quality of the annotation of regulatory interactions, or the absence of protein-metabolite interactions, and is also present when function is defined by Gene Ontology terms. Furthermore, the observed functional heterogeneity is different from the one expected by chance, supporting the notion that it is a biological property. To further explore the relationship between transcriptional regulation and metabolism, we analyze 5 other types of regulatory groups and identify complex regulons (i.e. genes regulated by the same combination of TFs) as the most functionally homogeneous, which is supported by coexpression data. Whether higher levels of related functions exist beyond metabolism and current functional annotations, remains an open question.