Effects of leucine-enkephalin and methionine-enkephalin on prolactin and gonadotropin secretion and synthesis in vitro

In vivo, Enkephalins, stimulate PRL, inhibit LH and are inactive on FSH. However, in monolayer pituitary cell cultures, PRL, LH and FSH secretions and synthesis are not modified by Met-Enk. (5 microgram/ml) or Leu-Enk. (5 and 10 microgram/ml). But the simultaneous presence of LHRH and Enk. induces an increase in LH secretion and synthesis without modifying FSH and PRL. In conclusion 1) Enk do not act by themself at the pituitary level but 2) they are able to modify the responses induced by hypothalamic hormones.     

Evidence against a role for alkaline phosphatase in the dephosphorylation of plasma membrane proteins: Hypophosphatasia fibroblast study

A major impasse to understanding the physiologic role(s) of alkaline phosphatase (ALP) is uncertainty as to its natural substrates. Various in vitro studies have led other investigators to suggest that ALP functions as a plasma membrane phosphoprotein phosphatase, consistent with our demonstration of ecto-topography of ALP in a variety of cell types. Thus, we compared the phosphorylation of plasma membrane proteins from control fibroblasts to those from profoundly ALP-deficient fibroblasts of hypophosphatasia patients. Fibroblasts from 3 controls and 3 hypophosphatasia patients (ALP activity < 4% of control) were biosynthetically labeled with ³²P_i for 2 h. ³²P incorporation into total trichloracetic acid (TCA)-precipitable material was not significantly different in control and patient cells. Plasma membranes were prepared from these cells by hypotonic shock, solubilized, and subjected to two-dimensional (2-D) gel electrophoretic separation. Video densitometric analysis of silver-stained 2-D gels failed to reveal any consistent difference in the protein profile between patient vs. control fibroblasts (i.e., unique species, altered pls, or increased abundance). Autoradiography of individual 2-D gels demonstrated 63 plasma membrane phosphoproteins with molecular weights ranging from 15 to 152 kDa and predominantly acidic pls. Although several of these phosphoproteins appeared to have had donor-specific labeling, none was unique or especially abundant in the hypophosphatasia group. Thus, in ALP-deficient fibroblasts, normal incorporation of ³²P into total cellular protein and into all identifiable plasma membrane phosphoproteins indicates that ALP does not modulate the phosphorylation of plasma membrane proteins.     

Substance use and other psychiatric disorders among 100 American Indian patients

One hundred American Indian patients with a Psychoactive Substance Use Disorder (PSUD) were studied with special reference to associated psychiatric disorders. This clinical sample was divided into three groups: PSUD only, PSUD plus an Organic Mental Disorder (OMD), and PSUD plus any other psychiatric disorder. OMD diagnoses included primarily Delirium Tremens and Alcoholic Hallucinosis; cases of Alcohol Amnestic Disorder, Alcohol Dementia, and trauma-induced OMD were also encountered. Other psychiatric disorders included primarily Major Depression and Anxiety Disorder, with smaller numbers of Schizophrenia, Conduct, Sexual, and other Disorders. Demographic and clinical characteristics were compared among these three groups. Those with PSUD + OMD tended to be older, male, and have more DSM-III Axis 3 disorders (American Psychiatric Association 1980) as compared to other patients; those with PSUD + other diagnoses tended to be single and younger. Education and occupational status were not related to the three diagnostic groups. The data were also subjected to MANOVA analysis. Even when corrected for sex, types of substance being abused, Axis 3 health status, and other factors, the three diagnostic groups still bore a significant relationship to age. Those with PSUD + Other psychiatric diagnoses besides OMD tended to be youngest. Those with PSUD-only were intermediate by age, while those with PSUD + OMD tended to be the oldest.     

Enthesis bilateral asymmetry in humans and African apes

Entheses (skeletal muscle and tendon attachment sites) have often been used to infer handedness and activity variability among human populations. However, the specific roles that intensity vs. frequency of muscle contractions play in modifying entheses are not well understood and the assumption that entheses reflect muscle activity levels has been challenged. This study explores the effect of habitual muscular activity on enthesis morphology in humans and African apes by investigating bilateral asymmetry in the forelimbs and hindlimbs of these taxa. Humans have generally more developed entheses in the lower limb while African apes have generally more developed entheses in the forelimbs. All species studied have more asymmetric forelimbs than hindlimbs except humans that show more asymmetrical expression of bony spurs in the lower limbs than in the upper limbs. When comparing species, humans are always more asymmetric in ethesis development than apes for both the forelimbs and hindlimbs, which reflects the relatively greater asymmetry in limb use in humans and the more symmetric use in apes. Enthesis development may reflect cross-symmetry patterns in humans and, more subtly, a moderate handedness in apes during manipulative activities. This study suggests that enthesis morphology provides information on muscle activity levels, with greater development of entheses associated with more habitual or powerful muscle use. The general similarity of ape and human responses to muscle activity suggests that muscle activity influenced enthesis development in Plio-Pleistocene hominins and that interpretation of muscle markings in these fossils can provide data for functional inferences in these extinct species.     

Snf1 Protein Kinase Regulates Phosphorylation of the Mig1 Repressor in Saccharomyces cerevisiae

In glucose-grown cells, the Mig1 DNA-binding protein recruits the Ssn6-Tup1 corepressor to glucose-repressed promoters in the yeast Saccharomyces cerevisiae. Previous work showed that Mig1 is differentially phosphorylated in response to glucose. Here we examine the role of Mig1 in regulating repression and the role of the Snf1 protein kinase in regulating Mig1 function. Immunoblot analysis of Mig1 protein from a snf1 mutant showed that Snf1 is required for the phosphorylation of Mig1; moreover, hxk2 and reg1 mutations, which relieve glucose inhibition of Snf1, correspondingly affect phosphorylation of Mig1. We show that Snf1 and Mig1 interact in the two-hybrid system and also coimmunoprecipitate from cell extracts, indicating that the two proteins interact in vivo. In immune complex assays of Snf1, coprecipitating Mig1 is phosphorylated in a Snf1-dependent reaction. Mutation of four putative Snf1 recognition sites in Mig1 eliminated most of the differential phosphorylation of Mig1 in response to glucose in vivo and improved the two-hybrid interaction with Snf1. These studies, together with previous genetic findings, indicate that the Snf1 protein kinase regulates phosphorylation of Mig1 in response to glucose.     

Directed evolution reveals the mechanism of HitRS signaling transduction in Bacillus anthracis

Two component systems (TCSs) are a primary mechanism of signal sensing and response in bacteria. Systematic characterization of an entire TCS could provide a mechanistic understanding of these important signal transduction systems. Here, genetic selections were employed to dissect the molecular basis of signal transduction by the HitRS system that detects cell envelope stress in the pathogen Bacillus anthracis. Numerous point mutations were isolated within HitRS, 17 of which were in a 50-residue HAMP domain. Mutational analysis revealed the importance of hydrophobic interactions within the HAMP domain and highlighted its essentiality in TCS signaling. In addition, these data defined residues critical for activities intrinsic to HitRS, uncovered specific interactions among individual domains and between the two signaling proteins, and revealed that phosphotransfer is the rate-limiting step for signal transduction. Furthermore, this study establishes the use of unbiased genetic selections to study TCS signaling and provides a comprehensive mechanistic understanding of an entire TCS.