Gene regulatory networks consist of direct interactions but also include indirect interactions mediated by metabolites and signaling molecules. We describe how these indirect interactions can be derived from a model of the underlying biochemical reaction network, using weak time-scale assumptions in combination with sensitivity criteria from metabolic control analysis. We apply this approach to a model of the carbon assimilation network in Escherichia coli. Our results show that the derived gene regulatory network is densely connected, contrary to what is usually assumed. Moreover, the network is largely sign-determined, meaning that the signs of the indirect interactions are fixed by the flux directions of biochemical reactions, independently of specific parameter values and rate laws. An inversion of the fluxes following a change in growth conditions may affect the signs of the indirect interactions though. This leads to a feedback structure that is at the same time robust to changes in the kinetic properties of enzymes and that has the flexibility to accommodate radical changes in the environment.

Alcaligenes eutrophus CH34 used benzoate as a sole source of carbon and energy, degrading it through the 3-oxoadipate pathway. All the enzymes required for this degradation were shown to be encoded by chromosomal genes. Catechol 1,2-dioxygenase activity was induced by benzoate, catechol, 4-chlorocatechol, and muconate. The enzyme is most likely a homodimer, with an apparent molecular weight of 76,000 +/- 500. According to several criteria, its properties are intermediate between those of catechol 1,2-dioxygenases (CatA) and chlorocatechol 1,2-dioxygenases (ClcA). The determined Km for catechol is the lowest among known catechol and chlorocatechol dioxygenases. Similar Km values were found for para-substituted catechols, although the catalytic constants were much lower. The catechol 1,2-dioxygenase from strain CH34 is unique in its property to transform tetrachlorocatechol; however, excess substrate led to a marked reversible inhibition. Some meta- and multi-substituted catechols behaved similarly. The determined Km (or Ki) values for para- or meta-substituted catechols suggest that the presence of an electron-withdrawing substituent at one of these positions results in a higher affinity of the enzyme for the ligand. Results of studies of recognition by the enzyme of various nonmetabolised aromatic compounds are also discussed.

A phylogenetic tree has been constructed from comparisons of entire 16S rRNA gene sequences from different prokaryotes and from several algal plastids. According to this study, and to previous work on the ribulose-1,5-bisphosphate carboxylase oxygenase (Rubisco) large and small subunit genes, we postulate that: (1) rhodophyte and chromophyte plastid genomes have a common, composite phylogenetic origin which implies at least two different ancestors, a cyanobacterial and a beta-proteobacterial ancestor; (2) chlorophyte (green algae and land plants) plastids have a cyanobacterial ancestor which probably differs from that of rhodophyte and chromophyte plastids, and in any case constitute a different lineage; (3) euglenophyte plastid genomes also seem to have a composite phylogenetic origin which involves two different lineages.

The algae have long been classified into three main groups based on their plastidial pigments (Christensen, 1964). In this scheme all three groups have the same primary pigment, chlorophyll a, and the different taxa are defined by their secondary pigments; the green algae having chlorophyll b, the red algae having phycobilins, and the chromophytes (which include the brown algae), having chlorophyll c. Today, plastidial characters are still viewed as important in defining the relationships among the algae, and between the algae and higher plants. In this light, some of the most intriguing questions about the chromophytes concern the different morphologies, origins, and evolutionary histories of their plastids. One way to study these organelles is to examine them at the genomic level and to compare both the gross organization and fine structure of their genes with those of other plastids (not only green plastids) and prokaryotes (not only cyanobacteria). For this purpose, we have begun an extensive investigation of the plastidial genome of a simple brown alga, Pylaiella littoralis (L.) Kjellm. Herein we describe progress to date in characterizing the ribosomal operons, genes and pseudogenes found on this genome, and we discuss the phylogenetic implications of these results.

The plastid genome of the brown alga Pylaiella littoralis (L.) Kjellm. is composed of two different circular DNA molecules: the largest carries two rrn operons, and the smallest, only one copy of both 16S and 23S rDNAs. 16S rDNA copies located on both molecules have been cloned and their nucleotide sequences determined: they are 65% homologous to one another. The expression of these genes was assayed by hybridizing in vivo labelled P. littoralis rRNAs to both clones, and specific oligonucleotides to total RNA from P. littoralis. Results indicate that the 16S rDNA copy located on the small molecule is a pseudogene. Comparisons of the functional gene with other 16S rRNA genes shows that chloroplasts from green plants emerged earlier from the cyanobacterial lineage than Euglena gracilis and Pylaiella littoralis plastids.

Two circular molecules of different sizes, both belonging to the chloroplast DNA of the brown algaPylaiella littoralis, have been observed by electron microscopy (Dalmon et al. 1983). Clone banks representing 86% of the small chloroplast circular DNA molecule (58 kbp) and 69% of the large circular DNA molecule (133 kbp) have been established and used as tools in the construction of physical maps. Two rDNA operons have been mapped in a very small inverted repeat on the large circular molecule. One 16S rRNA pseudogene and one split 23S rRNA gene have been mapped on the small DNA molecule, far apart from each other. Using heterologous probes, genes for ten different proteins have also been located on these maps. Their arrangement on the large molecule is different from that found in higher plants and algae. Probes fromrbcL, psbA andrps19 genes hybridize to several separated fragments. Two of them (psbA andrps19) hybridize to both types of molecules.

The DNA segment situated between the 16S and 23S rRNA genes belonging to the plastid genome of the brown alga Pylaiella littoralis (L.) Kjellm. has been sequenced. This small region (322 bp) contains two unsplit tRNA genes separated by 3 bp. A comparison with similar regions from different plants shows that this region has evolved in two different ways according to the place of plants in evolution. In the "primitive" group, this region is reduced in size when compared to prokaryotes. In the other groups, it is considerably enlarged by insertion of repetitive sequences, open reading frames and introns.