Microbiology, Adaptation and Pathogenesis Lab

In Kluyveromyces lactis, the expression of the major glucose permease gene RAG1 is controlled by extracellular glucose through a signaling cascade similar to the Saccharomyces cerevisiae Snf3/Rgt2/Rgt1 pathway. We have identified a key component of the K. lactis glucose signaling pathway by characterizing a new mutation, rag20-1, which impairs the regulation of RAG1 and hexokinase RAG5 genes by glucose. Functional complementation of the rag20-1 mutation identified the KlSNF2 gene, which encodes a protein 59% identical to S. cerevisiae Snf2, the major subunit of the SWI/SNF chromatin remodeling complex. Reverse transcription-quantitative PCR and chromatin immunoprecipitation analyses confirmed that the KlSnf2 protein binds to RAG1 and RAG5 promoters and promotes the recruitment of the basic helix-loop-helix Sck1 activator. Besides this transcriptional effect, KlSnf2 is also implicated in the glucose signaling pathway by controlling Sms1 and KlRgt1 posttranscriptional modifications. When KlSnf2 is absent, Sms1 is not degraded in the presence of glucose, leading to constitutive RAG1 gene repression by KlRgt1. Our work points out the crucial role played by KlSnf2 in the regulation of glucose transport and metabolism in K. lactis, notably, by suggesting a link between chromatin remodeling and the glucose signaling pathway.

A simple, rapid SDS-Page method for protein profiles analysis of Actinomyces species was developed. Eighteen strains (12 reference strains and 6 fresh isolates) representing four species were used in this study. Eight detergents were tested for protein extraction. Cell extracts were obtained by shaking the bacteria suspended in a solution containing a detergent and glass beads. After electrophoresis, according to the Laemmli's technique, two protein stain procedure were tested (Coomassie blue and silver stain). Best results were obtained with 0.5% Triton X-100 for 4 h and the silver stain procedure of Oakley. Protein profiles analysis showed that the method is reproducible and distinguishes not only species but sometimes also subspecies.

In many species, in vitro culture of fertilized embryos does not lead to normal cleavage and development as observed in vivo. When cultured in vitro, mouse eggs from outbred strains are blocked at the 2-cell stage unless they are transferred back into oviducts (Whittingham and Biggers, 1967). Hamster eggs also become blocked at this stage (Whittingham and Bavister, 1974) [see also Chapter 11 (Ed.)]. Most early bovine embryos (1- to 8-cell stage) arrest their development in vitro at the 8- to 12-cell stage (Thibault, 1966), in spite of a variety of culture media and conditions tested by several authors (reviewed by Wright and Bondioli, 1981).

The nutrition, growth, development and mortality of the cotton leafworm, Spodoptera littoralis, were followed on six different artificial diets. As compared with the reference diet of Poitout et al. (1972) (Entomologia exp. appl. 15, 341-350), our new media exhibit some interesting technical features: Different conditioning processes have been tested and this work was carried out to ensure that the two Korano gelling powders have the same biological quality as agar. The prepacked diets have proven to be very easy to store and to handle, and overall they are quite efficient and time-saving. Furthermore, they are much cheaper than the laboratory-prepared control diet. They supported the complete and normal development of S. littoralis, and even at a slightly better rate than agar-containing diets (M1 and M2b diets).