Supplementary materials for the paper:

“Reconstruction of regulatory and metabolic pathways in metal-reducing delta-proteobacteria”

Dmitry A. Rodionov*¹, Inna Dubchak², Adam Arkin³, Eric Alm³, and Mikhail S. Gelfand^1,4

¹Institute for Information Transmission Problems, Russian Academy of Sciences, Bolshoi Karetny per 19, Moscow, 127994, Russia;

²Genomics and ³Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, US;

⁴State Scientific Center GosNIIGenetika,1st Dorozhny pr. 1, Moscow 117545, Russia

* e-mail: rodionov@iitp.ru

ABSTRACT

Relatively little is known about the genetic basis for the unique physiology of metal-reducing genera in the delta subgroup of the proteobacteria. Recent availability of complete finished or draft quality genome sequence for seven representatives allowed us to investigate the genetic and regulatory factors in a number of key pathways involved in the biosynthesis of building blocks and cofactors, metal ion homeostasis, stress response, and energy metabolism using a combination of the regulatory sequence detection and analysis of the genomic context.

In the genomes of delta-proteobacteria, we identified candidate binding sites for four regulators of known specificity (BirA, CooA, HrcA, sigma-32), four types of metabolite-binding riboswitches (RFN, THI, B12 and S-box), and new binding sites for the FUR, ModE, NikR, PerR, and ZUR transcription factors, as well as for the previously uncharacterized factors HcpR and LysX. After reconstruction of the corresponding metabolic pathways and regulatory interactions, we identified possible functions for a large number of previously uncharacterized genes covering a wide range of cellular functions.

Phylogenetically diverse delta-proteobacteria appear to share their regulatory components with other bacteria. This study for the first time shows adaptability of comparative genomics approach for the de novo reconstruction of a regulatory network in poorly studied taxonomic groups of bacteria. Recent efforts in large-scale functional genomic characterization of Desulfovibrio species will provide a unique opportunity to test and expand our predictions.

A. Regulation mediated by DNA-binding transcription factors

1a. Biotin regulon (BirA)

2a. Lysine regulon (LysX)

3a. Ferric uptake regulon (FUR)

4a. Nickel uptake regulon (NikR)

5a. Zinc uptake regulon (ZUR)

6a. Molybdenum uptake regulon (ModE)

7a. Peroxide and oxidative stress regulon (PerR)

8a. Heat shock regulons (HrcA or CIRCE and s³²)

9a. Two FNR/CRP-like factors, the CO-responsive regulator CooA and a novel factor HcpR regulating energy metabolism

B. Regulation mediated by effector-binding RNA structural elements (riboswitches)

1b. Riboflavin biosynthesis (RFN element)

2b. Vitamin B12 biosynthesis (B12 element)

3b. Thiamin biosynthesis (THI element)

4b. Methionine biosynthesis (S-box)

P.S. Numerical gene identifiers from the Virtual Institute for Microbial Stress and Survival (VIMSS) Comparative Genomics database (http://escalante.lbl.gov/) are used throughout in the tables and figures.

C. Detailed description of software for genome analysis developed in the lab