Hoxc12 expression pattern in developing and cycling murine hair follicles

We examine the Hoxc12 RNA expression pattern during both hair follicle morphogenesis and cycling in direct comparison to its only upstream neighbor, Hoxc13. Expression of both genes is restricted to the epidermal part of the follicle excluding the outer root sheath and interfollicular epidermis in a distinct stage-dependent and cyclical manner. During the progressive growth phase (anagen) of developing and cycling follicles, the distinct proximo-distal expression domain of Hoxc12 overlaps only proximally, at the upper-most region of the bulb, with the more proximally restricted Hoxc13 domain. This arrangement of the expression domains of the two genes along the proximal-toward-distal axis of increasing follicular differentiation correlates with the sequential expression of first Hoxc13 and then Hoxc12. This indicates a reversal of the typical temporal colinearity of Hox gene activation otherwise observed along the anterior-posterior morphogenetic axis of the embryo (review: Cell 78 (1994) 191).     

Transcriptome analysis reveals a role of interferon-gamma in human neointima formation

The most effective immediate cure for coronary stenosis is stent-supported angioplasty. Restenosis due to neointima proliferation represents a major limitation. We investigated the expression of 2435 genes in atherectomy specimens and blood cells of patients with restenosis, normal coronary artery specimens, and cultured human smooth muscle cells (SMCs). Of the 223 differentially expressed genes, 37 genes indicated activation of interferon-gamma (IFN-gamma) signaling in neointimal SMCs. In cultured SMCs, IFN-gamma inhibited apoptosis. Genetic disruption of IFN-gamma signaling in a mouse model of restenosis significantly reduced the vascular proliferative response. Our data suggest an important role of IFN-gamma in the control of neointima proliferation.     

Functional analysis of heterologous GPCR signalling pathways in yeast

G protein-coupled receptors (GPCRs) regulate diverse biological processes in eukaryotes and such conservation allows an almost unrestricted interchange of signalling components between different cell types. Yeasts are attractive hosts in which to study GPCRs--they are amenable to both genetic and biochemical manipulation and their robustness, low cost and our ability to create strains that lack endogenous GPCRs make them ideal starting points for the development of assays suitable for high-throughput screening. Here we introduce readers to the possibilities of using yeast to analyse GPCRs describing the endogenous signalling pathways, the development of assays for heterologous GPCRs and the technology to elucidate GPCR structure and activity, focusing on the budding yeast Saccharomyces cerevisiae and recent developments using the fission yeast Schizosaccharomyces pombe.     

Lipid signalling pathways in normal and ras-transfected NIH/3T3 cells

The role of ras oncogenes in cellular signalling pathways involving phospholipid breakdown was studied in untransfected and proto-H-ras and mutated H-, K- and N-ras transfected NIH/3T3 cells. When the cells were grown at low cell densities, all of the ras transfected cells had 2-4 fold higher diacylglycerol (DAG) levels compared to growing NIH/3T3 cells. At high cell densities, DAG levels decreased in the former and increased in contact inhibited NIH/3T3 cells. In this regard, only cells transformed by mutated cellular and viral H-ras oncogenes (but not by the H-ras proto-oncogene) had elevated DAG levels compared to contact inhibited NIH/3T3 cells. The basal levels of inositol phosphates in ras transfected cells were not significantly different from NIH/3T3 cells and did not vary with cell density. Thus, the elevated DAG levels are not a consequence of increased phosphoinositide hydrolysis. The latter was stimulated by serum and bombesin only in normal and proto-H-ras transfected cells. In contrast, stimulation by bradykinin was observed only in cells transformed by mutated cellular ras oncogenes. Furthermore, aluminum fluoride stimulated phosphoinositide breakdown in the latter cells indicating that there was no uncoupling of the G protein from phospholipase C. Treatment of ras transfected cells with dibutyryl cyclic AMP (DB-cAMP), which causes an inhibition of growth and a reversal of the transformed morphology, did not alter the basal levels of inositol phosphates, DB-cAMP, however, did lower DAG levels in some of the transformed cell lines, but elevated DAG levels in low density NIH/3T3 cells. These findings indicate that the ras gene product p21 is not involved in phosphoinositide hydrolysis and that DAG levels do not correlate with cell growth in either normal or ras transfected NIH/3T3 cells. Thus, p21 appears to alter cell growth through mechanism(s) independent of lipid signalling pathways.     

Transcriptome analysis reveals absence of unintended effects in drought-tolerant transgenic plants overexpressing the transcription factor ABF3

Background

The superfamily of ABC proteins is among the largest known in nature. Its members are mainly, but not exclusively, involved in the transport of a broad range of substrates across biological membranes. Many contribute to multidrug resistance in microbial pathogens and cancer cells. The diversity of ABC proteins in fungi is comparable with those in multicellular animals, but so far fungal ABC proteins have barely been studied.

Results

We performed a phylogenetic analysis of the ABC proteins extracted from the genomes of 27 fungal species from 18 orders representing 5 fungal phyla thereby covering the most important groups. Our analysis demonstrated that some of the subfamilies of ABC proteins remained highly conserved in fungi, while others have undergone a remarkable group-specific diversification. Members of the various fungal phyla also differed significantly in the number of ABC proteins found in their genomes, which is especially reduced in the yeast S. cerevisiae and S. pombe.

Conclusions

Data obtained during our analysis should contribute to a better understanding of the diversity of the fungal ABC proteins and provide important clues about their possible biological functions.     

The human cysteine protease cathepsin V can compensate for murine cathepsin L in mouse epidermis and hair follicles

Mice lacking the ubiquitously expressed lysosomal cysteine protease cathepsin L, show a complex skin phenotype consisting of periodic hair loss and epidermal hyperplasia with hyperproliferation of basal epidermal keratinocytes, acanthosis and hyperkeratosis. The recently identified human cathepsin L-like enzyme cathepsin V, which is also termed cathepsin L2, is specifically expressed in cornea, testis, thymus, and epidermis. To date, in mice no cathepsin V orthologue with this typical expression pattern has been identified. Since cathepsin V has about 75% protein sequence identity to murine cathepsin L, we hypothesized that transgenic, keratinocyte-specific expression of cathepsin V in cathepsin L knockout mice might rescue the skin and hair phenotype. Thus, we generated a transgenic mouse line expressing cathepsin V under the control of the human keratin 14 promoter, which mimics the genuine cathepsin V expression pattern in human skin, by directing it to basal epidermal keratinocytes and the outer root sheath of hair follicles. Subsequently, transgenic mice were crossed with congenic cathepsin L knockout animals. The resulting mice show normalization of epidermal proliferation and normal epidermal thickness as well as rescue of the hair phenotype. These findings provide evidence for keratinocyte-specific pivotal functions of cathepsin L-like proteolytic activities in maintenance of epidermis and hair follicles and suggest, that cathepsin V may perform similar functions in human skin.