Alpha 1-antitrypsin and serum albumin mRNA accumulation in normal, acute phase and ZZ human liver

Alpha 1-Antitrypsin and albumin mRNA levels of 4 human livers were assessed using a newly sequenced cDNA clone of the carboxyterminal third of alpha 1-antitrypsin and a previously cloned albumin cDNA sequence. The relative concentration of alpha 1-antitrypsin mRNA was the same in poly(A)-containing RNA isolated from acute phase (MM) and alpha1-antitrypsin deficient (ZZ) individuals. In the acute phase liver relative to the normal (MM) liver, total RNA extracts showed a marked decrease in albumin mRNA concentration but no increase in alpha 1-antitrypsin mRNA. The ZZ liver showed decreased total and poly(A)-containing RNA content but the same proportion of alpha 1-antitrypsin to albumin mRNA as in the normal (MM) liver. This supports other evidence that ZZ alpha 1-antitrypsin deficiency is due to a defect in polypeptide processing (secretion) rather than a deficiency in mRNA accumulation.     

Arthrin: a new actin-like protein in insect flight muscle

There are one or more proteins of 50,000 to 60,000 Mr in the thin filaments of insect flight muscle. A protein of 55,000 Mr has been isolated from insect fibrillar flight muscle and called arthrin. Despite its higher molecular weight, arthrin is in many ways like actin. The amino acid composition of arthrin was similar to that of actin. There were similarities in the peptides produced by digesting the denatured proteins and mild digestion of polymerized proteins cleaved similar-sized fragments from arthrin and actin. Polymerized arthrin activated the Mg2+ ATPase of myosin to the same extent as actin and the ATPase was regulated by rabbit or Lethocerus troponin and tropomyosin. Arthrin did not itself act as troponin-T. Electron microscopy of negatively stained specimens showed that arthrin and actin filaments were similar in structure and that arthrin could be decorated by rabbit subfragment-1 to form normal-looking arrowheads. Arthrin formed paracrystals at an optimum concentration of MgCl2 (25 mM) that was somewhat lower than the optimum for actin paracrystals. Optical diffraction showed that the structure of the paracrystals was similar to those formed from actin. The mass of arthrin and actin filaments relative to phage fd was measured by scanning transmission electron microscopy; the relative mass of arthrin and actin was 1.33, in agreement with molecular weight estimations. Therefore arthrin has the properties of a heavy form of actin. The proportion of actin, arthrin and troponin-T in Lethocerus myofibrils was six moles of actin to one mole of arthrin and one mole of troponin-T. The function of arthrin is not known.     

Isolation and characterization of an alphoid centromeric repeat family from the human Y chromosome

A collection of human Y-derived cosmid clones was screened with a plasmid insert containing a member of the human X chromosome alphoid repeat family, DXZ1. Two positive cosmids were isolated and the repeats they contained were investigated by Southern blotting, in situ hybridization and sequence analysis. On hybridization to human genomic DNAs, the expected cross-hybridization characteristic of all alphoid sequences was seen and, in addition, a 5500 base EcoRI fragment was found to be characteristic of a Y-specific alphoid repeat. Dosage experiments demonstrated that there are about 100 copies of this 5500 base EcoRI alphoid fragment on the Y chromosome. Studies utilizing DNA from human-mouse hybrids containing only portions of the Y chromosome and in situ hybridizations to chromosome spreads demonstrated the Y centromeric localization of the 5500 base repeat. Cross-hybridization to autosomes 13, 14 and 15 was also seen; however, these chromosomes lacked detectable copies of the 5500 base EcoRI repeat sequence arrangement. Sequence analysis of portions of the Y repeat and portions of the DXZ1 repeat demonstrated about 70% homology to each other and of each to the human consensus alphoid sequence. The 5500 base EcoRI fragment was not seen in gorilla, orangutan or chimpanzee male DNA.     

5S and 5.8S ribosomal RNA evolution in the suborder tetrahymenina (Ciliophora: Hymenostomatida)

Summary The nucleotide sequences of the 5S and 5.8S rRNAs of eight strains of tetrahymenine ciliates have been determined. The sequences indicate a clear distinction betweenTetrahymena paravorax and its suggested conspecificT. vorax, but leave the taxonomic distinction betweenT. vorax andT. leucophrys in doubt. The rRNA sequences of sixTetrahymena species and of three other species of the suborder Tetrahymenina have been used to deduce evolutionary schemes in which ancestral rRNA sequences and changes are proposed. These schemes suggest the predominant acceptance of GA and CT transitions in the 5S rDNA during the evolution of the suborder.     

Cell-cell interaction can influence drug-induced differentiation of murine embryonal carcinoma cells

When cultured in the presence of either retinoic acid (RA) or dimethyl sulfoxide (DMSO), aggregates of the P19 line of mouse embryonal carcinoma (EC) cells differentiate and the spectrum of cell types formed depends on the drug dose. It is shown here the EC cells rapidly lose their colony-forming ability when cultured as aggregates in the presence of DMSO. This loss of plating efficiency (PE) also occurs rapidly following RA treatment. Loss of PE has been used as a quantitative procedure for assessing the rate of drug-induced differentiation. The relationship between drug dose and loss of PE is much steeper for DMSO than for RA, suggesting that these two drugs affect different stages of the differentiation decision-making apparatus. Mutant EC cell lines (D3 and RAC65) do not differentiate in the presence of drug-inducers (DMSO and RA, respectively). Neither differentiation-deficient mutant has an altered ability to form gap junctions. When D3 and P19 cells were mixed within the same DMSO-treated aggregates, the D3 cells remained undifferentiated and the P19 cells differentiated much less efficiently than if they were cultured in the absence of the D3 cells. When RAC65 and P19 cells were mixed in RA-treated aggregates, each cell responded to the drug as though the other were absent. Thus RA behaves as a cell-autonomous inducer of differentiation, whereas DMSO-induced differentiation seems to be mediated by interactions between neighboring cells.     

Studies of heterogeneous mitochondrial populations in a mouse cell line: the effects of selection for or against mitochondrial genomes that confer chloramphenicol resistance

A single nucleotide substitution in a highly conserved region of the mitochondrial genome of a mouse cell line confers both chloramphenicol resistance and an alteration to the recognition site for the endonuclease Eco RV. This has enabled a detailed study on the effects of selection on a mitochondrial population comprising initially both chloramphenicol-resistant and chloramphenicol-sensitive mitochondrial genomes. The mutation confers advantage to cells grown in the presence of chloramphenicol, but is apparently deleterious in its absence. Selection at the cellular level is sufficient to explain the results observed. Fixation, which results in cells having mitochondria of only a single type, is slow. It is probable, therefore, that mammalian oocyte mitochondria are derived from only a small number of progenitors. This would allow fixation of new mutations and explain the observed uniformity in mitochondrial genomes of the individual in the presence of extensive variation between different members of the population.     

Enzyme kinetics of a highly purified mitochondrial creatine kinase in comparison with cytosolic forms

Summary Mitochondrial creatine kinase (CK) purified from canine myocardium showed a single protein band on SDS-PAGE and was free of MMCK. Its amino acid composition was different than MMCK or BBCK and did not react to antiserum to MMCK or BBCK. Using purified mitochondrial, MM and BBCK, the velocity of reaction (V) was estimated for creatine phosphate (CP), creatine (C), adenosine triphosphate (ATP) and adenosine diphosphate (ADP) over a wide range of concentrations including those at V_max. The values for K_m (mM/L) derived from Lineweaver-Burke plots are shown: The affinity of mitochondrial CK for C is much greater than MMCK which is compatible with the energy shuttle hypothesis, namely ATP is converted by mitochondrial CK to CP, and then diffuses to the myofibril for conversion to ATP for utilization.     

Growth and mortality of zebra fish, Brachydanio rerio (Hamilton Buchanan), maintained at two temperatures and on two diets

One hundred zebra fish per tank were maintained for 112 days at 24°C or 28°C in glass aquaria and fed a diet of flaked food made without cellulose (13.45 kJ g^?1, metabolizable energy, Type A) or with cellulose (8.71 kJ g^?1, metabolizable energy, Type B). Each experimental condition was repeated in triplicate (12 tanks). The weight of food given daily to the fish was based on daily records of survivors (from which mortality rates were calculated) and wet wt of fish (measured every 14 days) in each tank. All fish were fed with the same weight of food per day and the quantity of energy in the food in excess of standard metabolism (as a proportion of SM) was approximately 0–5 for fish maintained at 28°C and fed food B, 1–0 for fish maintained at 24°C and fed food B, 1–5 for fish maintained at 28°C and fed food A, and 2.2 for fish maintained at 24°C and fed food A. Non-ionized ammonia, nitrite and nitrate nitrogen in the tanks did not reach toxic levels although there was an increase in total ammonium nitrogen in one tank and a subsequent heavy mortality. It was assumed that this was caused by the build up of pathogenic bacteria. Apart from this tank, mortality was highest in tanks at 28°C with fish fed food A and second highest in tanks at 24°C with fish fed on the same diet. Growth was measured in units of length, wet and dry weights, carbon and energy. There was a good correlation (P < 0.001) between carbon (mgC mg^?1) and calorific (J mg^?1) values and a conversion factor of 46.2 J (mgC)^?1 was derived. Fish maintained at 24°C and fed food A had the highest rates of growth both in weight and in energy value per unit weight. Fish fed the same diet but kept at 28°C had the lowest growth rates. Both these groups of fish had the highest coefficients of variation in wet weights which increased during the experiment, indicating an increase in interaction within the tanks. There was agreement between the energy value of fish sampled for growth and a condition factor based on the length-weight relationships of fish remaining in the tanks. A correlation (P < 0.05) was found between instantaneous mortality and growth rates for fish between tanks when those maintained at 28°C and fed on food A were ignored.