A plastome mutation affects processing of both chloroplast and nuclear DNA-encoded plastid proteins

Summary A bovine tRNA gene cluster has been characterized and the sequences of four tDNAs determined. Two of the tDNAs could encode tRNA^Ser _IGA, one tDNA^Ser _UGA, and the fourth tRNA^Gln _CUG. The three serine tDNAs representing the UCN codon isoacceptor family are almost identical. However, the sequence of the tDNA^Ser _TGA differs from a previously sequenced bovine tDNA^Ser _TGA at 12 positions (ca. 14%). This finding suggests that in the bovine genome, two subfamilies of genes might encode tRNA^Ser _UGA. It also raises the possibility that new genes for a specific UCN isoacceptor might arise from the genes of a different isoacceptor, and could explain previously observed differences between species in the anticodons of coevolving pairs of tRNAs^Ser _UCN. The gene cluster also contains complete and partial copies, and fragments, of the BCS (bovine consensus sequence) SINE (short interspersed nuclear element) family, six examples of which were sequenced. Some of these elements occur in close proximity to two of the serine tDNAs.     

Changes in the migratory properties of neural crest and early crest-derived cells in vivo following treatment with a phorbol ester drug

In previous work, we found that the phorbol ester drug 12-O-tetradecanoyl phorbol acetate (TPA) reversed the developmental restriction of melanogenesis that normally occurs in neural crest-derived Schwann cell precursors around embryonic Day 5 of quail development. That is, TPA treatment of dorsal root ganglia (DRG) from 7-day quail embryos caused Schwann cell precursors to regain the ability to give rise to melanocytes. In this paper, we examine other long-term effects of TPA on the differentiative and migratory properties of neural crest and crest-derived DRG cells, using heterospecific grafting methods. We report that TPA treatment in culture increased the extent of cell migration following grafting into host embryos, including some ectopic migration into the central nervous system and other locations. TPA did not, however, seem to change the fate of these crest-derived cells, except that some DRG cells underwent pigmentation, as had been observed previously. Interestingly, graft cells associated with peripheral nerves were found to be exclusively unpigmented, whereas graft cells found in all other locations, including the central nervous system, were both pigmented and unpigmented. This suggests that peripheral nerves may act in a fashion antagonistic to the effects of TPA. These findings are consistent with the notion that TPA treatment causes early crest-derived cells to regain developmental properties lost with developmental age.     

A calmodulin and alpha-subunit binding domain in human erythrocyte spectrin

Human erythrocyte spectrin binds calmodulin weakly under native conditions. This binding is enhanced in the presence of urea. The site responsible for this enhanced binding in urea has now been shown to reside in a specific region of the spectrin beta-subunit. Cleavage of spectrin with trypsin, cyanogen bromide or 2-nitro-5-thiocyanobenzoic acid generates fragments of the molecule which retain the ability to bind calmodulin under denaturing conditions. The origin of these fragments, identified by two-dimensional peptide mapping, is the terminal region of the spectrin beta-IV domain. The smallest peptide active in calmodulin binding is a 10 000 Mr fragment generated by cyanogen bromide cleavage. Only the intact 74 000 Mr fragment generated by trypsin (the complete beta-IV domain) retains the capacity to reassociate with the isolated alpha-subunit of spectrin. The position of a putative calmodulin binding site near a site for subunit-subunit association and protein 4.1 and actin binding suggests a possible role in vivo for calmodulin regulation of the spectrin-actin membrane skeleton or for regulation of subunit-subunit associations. This beta-subunit binding site in erythrocyte spectrin is found in a region near the NH2-terminus at a position analogous to the alpha-subunit calmodulin binding site previously identified in a non-erythroid spectrin by ultrastructural studies.     

Asthma mortality: comparison between New Zealand and England.

Causes for the high mortality from asthma in New Zealand were investigated by comparing deaths from asthma in caucasian subjects aged 15-64 in New Zealand with those from asthma in the same age group in two regions in England. There were no significant differences in the accuracy of death certification. The verified asthma mortality in New Zealand (4.2/100,000) was over twice that in England. Many characteristics of patients and management, including poor compliance with treatment and deficiencies in long term and emergency care, were qualitatively similar in the two countries. New Zealand had an apparently higher rate of non-preventable deaths from asthma, suggesting a greater severity of asthma in New Zealand. In both countries, however, most deaths were associated with poor assessment, underestimation of severity and inappropriate treatment (over-reliance on bronchodilators and underuse of systemic corticosteroids), and delays in obtaining help. A greater frequency of some of these deficiencies in management remains a possible additional explanation for part of the excess mortality in New Zealand.     

Organization and stability of endogenous xenotropic murine leukemia virus proviral DNA in mouse genomes.

In a series of blot hybridization experiments, using a xenotropic envelope probe and restriction enzymes known to cut xenotropic proviral DNA a single time (EcoRI) or not at all (HindIII), we have studied the organization and relationship of endogenous xenotropic env-related sequences in various mouse strains. Multiple copies (18 to 28) of xenotropic env-reactive fragments were found in all mouse DNAs after digestion with either HindIII or EcoRI, and the majority of fragments were of sizes compatible with their origin from full-length proviral DNA. Five HindIII and five EcoRI restriction fragments were common to all inbred mouse DNAs tested. In addition, each strain exhibited unique characteristic xenotropic env-reactive bands; these bands were remarkably stable during many years of inbreeding. The cleavage patterns characteristic of each strain were also useful for showing genealogical relatedness among the various inbred mice.     

Membrane composition and physiological activity of plastids from an oenothera plastome mutator-induced chloroplast mutant

Plastids were isolated from a plastome mutator-induced mutant (pm7) of Oenothera hookeri and were analyzed for various physiological and biochemical attributes. No photosynthetic electron transport activity was detected in the mutant plastids. This is consistent with previous ultrastructural analysis showing the absence of thylakoid membranes in the pm7 plastids and with the observation of aberrant processing and accumulation of chloroplast proteins in the mutant. In comparison to wild type, the mutant tissue lacks chlorophyll, and has significant differences in levels of four fatty acids. The analyses did not reveal any differences in carotenoid levels nor in the synthesis of several chloroplast lipids. The consequences of the altered composition of the chloroplast membrane are discussed in terms of their relation to the aberrant protein processing of the pm7 plastids. The pigment, fatty acid, and lipid measurements were also performed on two distinct nuclear genotypes (A/A and A/C) which differ in their compatibility with the plastid genome (type I) contained in these lines. In these cases, only chlorophyll concentrations differed significantly.     

Differentiation-dependent expression of the brown adipocyte uncoupling protein gene: regulation by peroxisome proliferator-activated receptor gamma.

Uncoupling protein (UCP) is expressed only in brown adipocytes and is responsible for the unique thermogenic properties of this cell type. The novel brown preadipocyte cell line, HIB-1B, expresses UCP in a strictly differentiation-dependent manner. Transgenic mice studies have shown that a region from kb -2.8 to -1.0 of the marine UCP gene is required for brown adipocyte-specific expression. Subsequent analysis identified a potent 220-bp enhancer from kb -2.5 to -2.3. We show that this enhancer is active only in differentiated HIB-1B adipocytes, and we identify a peroxisome proliferator-activated receptor gamma (PPARgamma) response element, referred to as UCP regulatory element 1 (URE1), within the enhancer. URE1 has differentiation-dependent enhancing activity in HIB-1B cells and is required for enhancer action, since mutations of URE1 that block protein binding abolish enhancer activity. We also show that PPAR gamma antibodies block binding to URE1 of nuclear extracts from cultured brown adipocytes and from the brown adipose tissue of cold-exposed mice. Protein binding to URE1 increases substantially during differentiation of HIB-1B preadipocytes, and PPAR-gamma mRNA levels increase correspondingly. Although forced expression of PPAR gamma and retinoid X receptor alpha activates the enhancer in HIB-1B preadipocytes, these receptors are not capable of activating the enhancer in NIH 3T3 fibroblasts. Our results show that PPAR gamma is a regulator of the differentiation-dependent expression of UCP and suggest that there are additional factors in HIB-1B cells required for brown adipocyte-specific UCP expression.