Variability of uterine cervical anatomy in the white-tailed gnu (Connochaetes gnou)

Examination of the reproductive anatomy of six female white-tailed gnu revealed duplex uterus with varying degrees of cervical division. The division ranged from 36 to 100% of the length of the cervix, with one animal exhibiting double external ossa. This variability in reproductive anatomy could be important in artificial insemination or embryo transfer procedures in this species.     

Expression and characterization of a functional human insulin-like growth factor I receptor

Stable transfectants of Chinese hamster ovary (CHO) cells were developed that expressed the protein encoded by a human insulin-like growth factor I (IGF-I) receptor cDNA. The transfected cells expressed approximately 25,000 high affinity receptors for IGF-I (apparent Kd of 1.5 X 10(-9) M), whereas the parental CHO cells expressed only 5,000 receptors per cell (apparent Kd of 1.3 X 10(-9) M). A monoclonal antibody specific for the human IGF-I receptor inhibited IGF-I binding to the expressed receptor and immunoprecipitated polypeptides of apparent Mr values approximately 135,000 and 95,000 from metabolically labeled lysates of the transfected cells but not control cells. The expressed receptor was also capable of binding IGF-II with high affinity (Kd approximately 3 nM) and weakly recognized insulin (with about 1% the potency of IGF-I). The human IGF-I receptor expressed in these cells was capable of IGF-I-stimulated autophosphorylation and phosphorylation of endogenous substrates in the intact cell. This receptor also mediated IGF-I-stimulated glucose uptake, glycogen synthesis, and DNA synthesis. The extent of these responses was comparable to the stimulation by insulin of the same biological responses in CHO cells expressing the human insulin receptor. These results indicate that the isolated cDNA encodes a functional IGF-I receptor and that there are no inherent differences in the abilities of the insulin and IGF-I receptors to mediate rapid and long term biological responses when expressed in the same cell type. The high affinity of this receptor for IGF-II also suggests that it may be important in mediating biological responses to IGF-II as well as IGF-I.     

Trimethyloxonium modification of single batrachotoxin-activated sodium channels in planar bilayers. Changes in unit conductance and in block by saxitoxin and calcium

Single batrachotoxin-activated sodium channels from rat brain were modified by trimethyloxonium (TMO) after incorporation in planar lipid bilayers. TMO modification eliminated saxitoxin (STX) sensitivity, reduced the single channel conductance by 37%, and reduced calcium block of inward sodium currents. These effects always occurred concomitantly, in an all-or-none fashion. Calcium and STX protected sodium channels from TMO modification with potencies similar to their affinities for block. Calcium inhibited STX binding to rat brain membrane vesicles and relieved toxin block of channels in bilayers, apparently by competing with STX for the toxin binding site. These results suggest that toxins, permeant cations, and blocking cations can interact with a common site on the sodium channel near the extracellular surface. It is likely that permeant cations transiently bind to this superficial site, as the first of several steps in passing inward through the channel.     

Enzymatic mechanism for the hydrolysis of 5,10-methenyltetrahydropteroylglutamate to 5-formyltetrahydropteroylglutamate by serine hydroxymethyltransferase.

Serine hydroxymethyltransferase in the presence of glycine catalyzes the hydrolysis of (6R)-5,10-methenyltetrahydropteroylpolyglutamate to (6S)-5-formyltetrahydropteroylpolyglutamate. The enzyme also catalyzes the formation of (6S)-5-formyltetrahydropteroylpolyglutamate from a compound in equilibrium with (6R)-5,10-methenyltetrahydropteroylpolyglutamate believed to be (6R,11R)-5,10-hydroxymethylenetetrahydropteroylpolyglutamate , a putative intermediate in the nonenzymatic hydrolysis of 5,10-methenyltetrahydropteroylglutamate to 5-formyltetrahydropteroylglutamate [Stover, P., & Schirch, V. (1992) Biochemistry (preceding paper in this issue)]. The enzymatic mechanism for the formation of (6S)-5-formyltetrahydropteroylpolyglutamate from these substrates and the role of glycine in the reaction was addressed. Evidence suggests that (6R,11R)-5,10-hydroxymethylenetetrahydropteroyltetraglutamate++ + is a catalytically competent intermediate in the enzyme-catalyzed hydrolysis of (6R)-5,10-methenyltetrahydropteroyltetraglutamate. The enzyme displays a high Km of 40 microM for (6R)-5,10-methenyltetrahydropteroyltetraglutamate, while the Km for (6R,11R)-5,10-hydroxymethylenetetrahydropteroyltetraglutamate++ + is below 0.5 microM. The kcat values for both reactions are identical and equal to the rate of formation of an enzyme ternary complex absorbing at 502 nm which is formed from glycine and (6S)-5-formyltetrahydropteroylpolyglutamate. The hydrolysis reaction proceeds with exchange of the C11 formyl proton of (6R)-5,10-methenyltetrahydropteroyltetraglutamate, suggesting that the enzyme-catalyzed reaction occurs by the same C11 carbanion inversion mechanism as the nonenzymatic reaction. Isotope exchange experiments using [2-3H]glycine and differential scanning calorimetry data suggest both a catalytic and a conformational role for glycine in the enzymatic reaction. The results are discussed in terms of the similarity in mechanisms of the SHMT-catalyzed retroaldol cleavage of serine and hydrolysis of (6R)-5,10-methenyltetrahydropteroylpolyglutamates.     

Haploinsufficiency of cytosolic serine hydroxymethyltransferase in the Smith-Magenis syndrome.

Folate-dependent one-carbon metabolism is critical for the synthesis of numerous cellular constituents required for cell growth, and serine hydroxymethyltransferase (SHMT) is central to this process. Our studies reveal that the gene for cytosolic SHMT (cSHMT) maps to the critical interval for Smith-Magenis syndrome (SMS) on chromosome 17p11.2. The basic organization of the cSHMT locus on chromosome 17 was determined and was found to be deleted in all 26 SMS patients examined by PCR, FISH, and/or Southern analysis. Furthermore, with respect to haploinsufficiency, cSHMT enzyme activity in patient lymphoblasts was determined to be approximately 50% that of unaffected parent lymphoblasts. Serine, glycine, and folate levels were also assessed in three SMS patients and were found to be within normal ranges. The possible effects of cSHMT hemizygosity on the SMS phenotype are discussed.     

Biological activities of melanotropic peptide fatty acid conjugates.

Four fatty acids (FA, palmitic, myristic, decanoic, hexanoic) were individually conjugated to the N-terminus of the alpha-MSH fragment analog, H-Asp5-His6-D-Phe7-Arg8-Trp9-Lys10-NH2. This resulted in enhanced potency of the conjugates (compared to the unconjugated melanotropin analog) as determined in the lizard skin bioassay and in the mouse melanoma cell tyrosinase bioassay. The shorter conjugates of hexanoic and decanoic acid were at least equipotent to alpha-MSH in the lizard skin bioassay, whereas the longer myristoyl and palmitoyl analogs were 100 times less active. The myristoyl and palmitoyl conjugates exhibited a "creeping" potency in the lizard skin bioassay-that is, potency of the peptides increased with time in contact with the skins. These observations may be related to the more lipid nature of these FA-conjugates. In the tyrosinase assay, the conjugates were 10-100 times more active than alpha-MSH or the unconjugated analog. Each of the FA-melanotropic peptide conjugates exhibited prolonged (residual) melanotropic activity in both the lizard skin and melanoma cell bioassays. In other words, after removal of the melanotropin conjugates from contact with the skins or cells, responses were still manifested for hours or days thereafter. As little as 1 hr of contact with melanoma cells resulted in enhanced enzyme activity as measured 48 hr later. Since the conjugates, but not H-[Asp5, D-Phe7, Lys10]alpha-MSH5-10-NH2, exhibited prolonged activity, the conversion of reversible agonists to irreversible agonists was demonstrated.     

Interactions of the receptor for insulin-like growth factor II with mannose-6-phosphate and antibodies to the mannose-6-phosphate receptor

Recently, the sequence of the human receptor for insulin-like growth factor II (IGF-II) was found to be 80% identical [Morgan et al., (1987) Nature 329, 301-307] to the sequence of a partial clone of the bovine cation-independent mannose-6-phosphate receptor [Lobel et al., (1987) Proc. Natl. Acad. Sci. USA 84, 2233-2237]. In the present study, the purified receptor for insulin-like growth factor II (IGF-II) was found to react with two different polyclonal antibodies to the purified mannose-6-phosphate receptor. Moreover, mannose-6-phosphate was found to stimulate the binding of labeled IGF-II to the IGF-II receptor by two-fold. This effect had the same specificity and affinity as the reported binding of mannose-6-phosphate to its receptor; mannose-1-phosphate and mannose had no effect on the binding of labeled IGF-II to its receptor, and the half-maximally effective concentration of mannose-6-phosphate was 0.3 mM. Also, mannose-6-phosphate did not affect labeled IGF-II binding to the insulin receptor. These results support the hypothesis that a single protein of Mr-250,000 binds both IGF-II and mannose-6-phosphate. Furthermore, they indicate that mannose-6-phosphate can modulate the interaction of IGF-II to its receptor.     

The Oxytricha trifallax Macronuclear Genome: A Complex Eukaryotic Genome with 16,000 Tiny Chromosomes

The macronuclear genome of the ciliate Oxytricha trifallax displays an extreme and unique eukaryotic genome architecture with extensive genomic variation. During sexual genome development, the expressed, somatic macronuclear genome is whittled down to the genic portion of a small fraction (∼5%) of its precursor “silent” germline micronuclear genome by a process of “unscrambling” and fragmentation. The tiny macronuclear “nanochromosomes” typically encode single, protein-coding genes (a small portion, 10%, encode 2–8 genes), have minimal noncoding regions, and are differentially amplified to an average of ∼2,000 copies. We report the high-quality genome assembly of ∼16,000 complete nanochromosomes (∼50 Mb haploid genome size) that vary from 469 bp to 66 kb long (mean ∼3.2 kb) and encode ∼18,500 genes. Alternative DNA fragmentation processes ∼10% of the nanochromosomes into multiple isoforms that usually encode complete genes. Nucleotide diversity in the macronucleus is very high (SNP heterozygosity is ∼4.0%), suggesting that Oxytricha trifallax may have one of the largest known effective population sizes of eukaryotes. Comparison to other ciliates with nonscrambled genomes and long macronuclear chromosomes (on the order of 100 kb) suggests several candidate proteins that could be involved in genome rearrangement, including domesticated MULE and IS1595-like DDE transposases. The assembly of the highly fragmented Oxytricha macronuclear genome is the first completed genome with such an unusual architecture. This genome sequence provides tantalizing glimpses into novel molecular biology and evolution. For example, Oxytricha maintains tens of millions of telomeres per cell and has also evolved an intriguing expansion of telomere end-binding proteins. In conjunction with the micronuclear genome in progress, the O. trifallax macronuclear genome will provide an invaluable resource for investigating programmed genome rearrangements, complementing studies of rearrangements arising during evolution and disease.     

Exploring the Potential Health Impact and Cost-Effectiveness of AIDS Vaccine within a Comprehensive HIV/AIDS Response in Low- and Middle-Income Countries

Background

The Investment Framework Enhanced (IFE) proposed in 2013 by the Joint United Nations Programme on HIV/AIDS (UNAIDS) explored how maximizing existing interventions and adding emerging prevention options, including a vaccine, could further reduce new HIV infections and AIDS-related deaths in low- and middle-income countries (LMICs). This article describes additional modeling which looks more closely at the potential health impact and cost-effectiveness of AIDS vaccination in LMICs as part of UNAIDS IFE.

Methods

An epidemiological model was used to explore the potential impact of AIDS vaccination in LMICs in combination with other interventions through 2070. Assumptions were based on perspectives from research, vaccination and public health experts, as well as observations from other HIV/AIDS interventions and vaccination programs. Sensitivity analyses varied vaccine efficacy, duration of protection, coverage, and cost.

Results

If UNAIDS IFE goals were fully achieved, new annual HIV infections in LMICs would decline from 2.0 million in 2014 to 550,000 in 2070. A 70% efficacious vaccine introduced in 2027 with three doses, strong uptake and five years of protection would reduce annual new infections by 44% over the first decade, by 65% the first 25 years and by 78% to 122,000 in 2070. Vaccine impact would be much greater if the assumptions in UNAIDS IFE were not fully achieved. An AIDS vaccine would be cost-effective within a wide range of scenarios.

Interpretation

Even a modestly effective vaccine could contribute strongly to a sustainable response to HIV/AIDS and be cost-effective, even with optimistic assumptions about other interventions. Higher efficacy would provide even greater impact and cost-effectiveness, and would support broader access. Vaccine efficacy and cost per regimen are critical in achieving cost-effectiveness, with cost per regimen being particularly critical in low-income countries and at lower efficacy levels.