The CXCL12γ Chemokine Displays Unprecedented Structural and Functional Properties that Make It a Paradigm of Chemoattractant Proteins

The CXCL12γ chemokine arises by alternative splicing from Cxcl12, an essential gene during development. This protein binds CXCR4 and displays an exceptional degree of conservation (99%) in mammals. CXCL12γ is formed by a protein core shared by all CXCL12 isoforms, extended by a highly cationic carboxy-terminal (C-ter) domain that encompass four overlapped BBXB heparan sulfate (HS)-binding motifs. We hypothesize that this unusual domain could critically determine the biological properties of CXCL12γ through its interaction to, and regulation by extracellular glycosaminoglycans (GAG) and HS in particular. By both RT-PCR and immunohistochemistry, we mapped the localization of CXCL12γ both in mouse and human tissues, where it showed discrete differential expression. As an unprecedented feature among chemokines, the secreted CXCL12γ strongly interacted with cell membrane GAG, thus remaining mostly adsorbed on the plasmatic membrane upon secretion. Affinity chromatography and surface plasmon resonance allowed us to determine for CXCL12γ one of the higher affinity for HS (K_d = 0.9 nM) ever reported for a protein. This property relies in the presence of four canonical HS-binding sites located at the C-ter domain but requires the collaboration of a HS-binding site located in the core of the protein. Interestingly, and despite reduced agonist potency on CXCR4, the sustained binding of CXCL12γ to HS enabled it to promote in vivo intraperitoneal leukocyte accumulation and angiogenesis in matrigel plugs with much higher efficiency than CXCL12α. In good agreement, mutant CXCL12γ chemokines selectively devoid of HS-binding capacity failed to promote in vivo significant cell recruitment. We conclude that CXCL12γ features unique structural and functional properties among chemokines which rely on the presence of a distinctive C-ter domain. The unsurpassed capacity to bind to HS on the extracellular matrix would make CXCL12γ the paradigm of haptotactic proteins, which regulate essential homeostatic functions by promoting directional migration and selective tissue homing of cells.     

Safety and Immunomodulatory Effects of Three Probiotic Strains Isolated from the Feces of Breast-Fed Infants in Healthy Adults: SETOPROB Study

We previously described the isolation and characterization of three probiotic strains from the feces of exclusively breast-fed newborn infants: Lactobacillus paracasei CNCM I-4034, Bifidobacterium breve CNCM I-4035 and Lactobacillus rhamnosus CNCM I-4036. These strains were shown to adhere to intestinal mucus in vitro, to be sensitive to antibiotics and to resist biliary salts and low pH. In the present study, a multicenter, randomized, double-blind, placebo-controlled trial with 100 healthy volunteers in three Spanish cities was carried out to evaluate the tolerance, safety, gut colonization and immunomodulatory effects of these three probiotics. Volunteers underwent a 15-day washout period, after which they were randomly divided into 5 groups that received daily a placebo, a capsule containing one of the 3 strains or a capsule containing a mixture of two strains for 30 days. The intervention was followed by another 15-day washout period. Patients did not consume fermented milk for the entire duration of the study. Gastrointestinal symptoms, defecation frequency and stool consistency were not altered by probiotic intake. No relevant changes in blood and serum, as well as no adverse events occurred during or after treatment. Probiotic administration slightly modified bacterial populations in the volunteers’ feces. Intestinal persistence occurred in volunteers who received L. rhamnosus CNCM I-4036. Administration of B. breve CNCM I-4035 resulted in a significant increase in fecal secretory IgA content. IL-4 and IL-10 increased, whereas IL-12 decreased in the serum of volunteers treated with any of the three strains. These results demonstrate that the consumption of these three bacterial strains was safe and exerted varying degrees of immunomodulatory effects.

Trial Registration

ClinicalTrials.gov NCT01479543     

Homeotic gene expression in the wild-type and a homeotic mutant of the moth Manduca sexta

Antibodies were used to examine the expression patterns of Antennapedia (Antp), Ultrabithorax (Ubx), Ubx and abdominal-A combined(Ubx/abd-A),and Distalless (Dll) in the embryos of the moth Manduca sexta. We found that the spatial and temporal pattern of Antp expression in Manduca was correlated with the anterior migration of two patches of epithelium that include the anterior-most tracheal pits, and with the development of functional spiracles. Ubx expression showed an intricate pattern which suggests complex regulation during development. Throughout Manduca embryogenesis the expression of Ubx/Abd-A and Dll was similar to that reported for other insects. However, there was no apparent reduction in Ubx/Abd-A expression in the Manduca abdominal proleg primordia that expressed Dll. The expression of these four proteins was also examined in embryosof the Manduca homozygous homeotic mutant Octopod (Octo). The Octo mutation results in the transformation of A1 and A2 in the anterior direction, with homeotic legs appearing on A1 and occasionally A2. Our results suggest that in Octo animals there is a reduction in the level of Ubx protein expression throughout its domain. Based on homeotic gene expression in wild-type and mutant Manduca and in other insects, we discuss potential roles of homeotic genes in insect morphological evolution. Received: 21 September 1998 / Accepted: 5 March 1999     

Activities of the protein kinases STK, PI3K, MEK, and ERK are required for the development of the head organizer in Hydra magnipapillata

The development of the hydra's head and its hypostome has been studied at the molecular level. Many genes have been cloned from hydra as potential candidates that control the development of its head. Much work was performed on the mechanisms controlling expression of these genes in the position-dependent manner. Moreover, there have been data to support the involvement of three main signaling pathways that involve PKC, SRC, and PI3K kinases in the regulation of the head formation and in the expression of several head-specific genes. In this report, we present data supporting the participation of these three signaling pathways on the development of the hypostome. We used grafting experiments and inhibitors of the specific kinases to show the participation of these enzymes in hypostome formation. From our results, we postulate that these signal transduction pathways regulate the very early stages of the head development, most likely at the point when the cells start to differentiate to form the head organizer.     

Bamboo flowers visited by insects: do insects play a role in the pollination of bamboo flowers?

Relatively little is known about pollination and other aspects of the reproductive biology of bamboos, but wind pollination is assumed to be the rule, at least in woody bamboos. Documenting the reproductive biology of woody bamboos is a complex task due to the long periods of time between flowering cycles, which range from 3 to 120 years. Insects visiting Guadua paniculata and G. inermis flowers were collected in the field. Scanning electron micrographs were taken of the visiting insects. Four species of bees, three from tribe Meliponini (Geotrigona acapulconis, Plebeia frontalis and Trigona fulviventris) and one from tribe Apini (Apis mellifera), along with a syrphid fly (Toxomerus teligera) were found visiting bamboo flowers. Some species of Hemiptera were also found feeding on the flowers, such as Neortholomus jamaicensis (Lygaeidae), or preying on the flower visitors (Apiomerus pictipes (Reduviidae)). Insects visiting bamboo inflorescences may facilitate the release of pollen grains into the air, promoting outcrossing and genetic flow among the individuals of the flowering bamboo populations.     

Comparative study of the sympatric ferns Culcita macrocarpa and Woodwardia radicans: Sexual phenotype

The sexual phenotypes of 1152 gametophytes from four populations of Culcita macrocarpa and Woodwardia radicans were monitored over a 1-year period. Gametophytes were maintained under three experimental conditions: (1) isolated, (2) pairs from the same sporophyte, or (3) pairs from different sporophytes. The frequencies of the sexual phenotypes did not vary significantly among these three conditions, and although there were some quantitative differences between populations, the sexual-phenotype sequences observed were species-specific. Gametophytes of C. macrocarpa were first male and then hermaphrodite: this sequence, together with the absence of antheridiogens, favours intragametophytic selfing. Natural populations of C. macrocarpa are presumably androdioecious. Gametophytes of W. radicans were first female and then hermaphrodite: this sequence and antheridiogen activity favour intergametophytic and even xenogamous mating. Despite these laboratory findings, populations of W. radicans are probably trioecious (because of the effects of antheridiogen). Few sporophytes of W. radicans were obtained in the present study, and none of C. macrocarpa: this is attributable to limiting illumination or substrate.     

A molecular biological protocol to distinguish potentially human pathogenic Stenotrophomonas maltophilia from plant-associated Stenotrophomonas rhizophila

In recent years, the importance of the Gram-negative bacterium Stenotrophomonas as an opportunistic pathogen as well as in biotechnology has increased. The aim of the present study was to develop new methods for distinguishing between strains closely related to the potentially human pathogenic Stenotrophomonas maltophilia and those closely related to the plant-associated Stenotrophomonas rhizophila. To accomplish this, 58 strains were characterized by 16S rDNA sequencing and amplified ribosomal DNA restriction analysis (ARDRA), and the occurrence of specific functional genes. Based on 16S rDNA sequences, an ARDRA protocol was developed which allowed differentiation between strains of the S. maltophilia and the S. rhizophila group. As it was known that only salt-treated cells of S. rhizophila were able to synthesize the compatible solute glucosylglycerol (GG), the ggpS gene responsible for GG synthesis was used for differentiation between both species and it was confirmed that it only occurred in S. rhizophila strains. As a further genetic marker the smeD gene, which is part of the genes coding for the multidrug efflux pump SmeDEF from S. maltophilia, was used. Based on the results we propose a combination of fingerprinting techniques using the 16S rDNA and the functional genes ggpS and smeD to distinguish both Stenotrophomonas species.     

Temporally regulated expression of Cre recombinase in neural stem cells

The use of mouse gene targeting to study molecules important in neural development is oftentimes impaired by early embryonic lethality. In order to address later roles for such molecules, specifically in neural stem cells, we generated transgenic mice that express both the tetracycline-inducible molecule rtTA-M2 and GFP under the control of the neural precursor specific form of nestin. Developmental analysis of these mice demonstrates that GFP expression is exclusive to the neural tube. Adult expression of GFP is seen only in known areas of adult neurogenesis, namely, the subventricular zone and the dentate gyrus. When crossed with a second transgenic mouse (TetOp-Cre) that expresses the Cre recombinase under the control of the tetracycline responsive promotor, we demonstrate temporal induction of Cre in bigenic animals exposed to doxycycline. We further demonstrate the feasibility of this approach by using the ROSA-26 reporter mouse to mediate recombination in neural precursor cells.     

Looking to the future with optimism: An interview with Glenn Nierman on the occasion of the recent end of his appointment as NAfME president

The author interviewed Glenn Nierman on the occasion of the recent conclusion of his term as National Association for Music Education (NAfME) president. Topics included the current state of music education and advocacy of music education in the United States, the role of NAfME, and the recently revised national music education standards.