Novel surface lipids of diapausing Manduca sexta pupae. Long chain oxoalcohol esters of acetoacetic, hydroxybutyric, and acetic acids

Ester components in the surface wax from diapausing tobacco hornworm pupae, Manduca sexta L., were separated by thin layer chromatography and gas-liquid chromatography, and characterized by infrared spectroscopy and gas-liquid chromatography-mass spectrometry. Three groups of esters were identified as natural derivatives of acetic acid, acetoacetic acid, and 3-hydroxybutyric acid. The major ester fraction was identified as a mixture of C26 (10%), C27 (5%), and C28 (85%) oxoalcohol esters of acetoacetic acid. The major homolog consisted of equal amounts of 11-oxooctacosanyl 3-oxobutanoate and 12-oxooctacosanyl 3-oxobutanoate. Lesser amounts of 11- and 12-oxooctacosanyl and n-octacosanyl esters of acetic and 3-hydroxybutyric acids were also identified. The chain length distributions of these C26, C27, and C28 oxoalcohol and n-primary alcohol ester moieties, as well as the isomeric ratios for the 11- and 12-oxoalcohol isomers, were similar to the oxoaldehydes and unesterified oxoalcohols previously identified by Buckner et al (Buckner, J. S., Nelson, D. R., Haak, H., and Pomonis, J. G. (1984) J. Biol. Chem. 259, 8452-8470) as lipid components of the surface wax of M. sexta pupae.     

Effect of flap modifications on human FEN1 cleavage.

The flap endonuclease, FEN1, plays a critical role in DNA replication and repair. Human FEN1 exhibits both a 5' to 3' exonucleolytic and a structure-specific endonucleolytic activity. On primer-template substrates containing an unannealed 5'-tail, or flap structure, FEN1 employs a unique mechanism to cleave at the point of annealing, releasing the 5'-tail intact. FEN1 appears to track along the full length of the flap from the 5'-end to the point of cleavage. Substrates containing structural modifications to the flap have been used to explore the mechanism of tracking. To determine whether the nuclease must recognize a succession of nucleotides on the flap, chemical linkers were used to replace an interior nucleotide. The nuclease could readily traverse this site. The footprint of the nuclease at the time of cleavage does not extend beyond 25 nucleotides on the flap. Eleven-nucleotide branches attached to the flap beyond the footprinted region do not prevent cleavage. Single- or double-thymine dimers also allow cleavage. cis-Platinum adducts outside the protected region are moderately inhibitory. Platinum-modified branch structures are completely inert to cleavage. These results show that some flap modifications can prevent or inhibit tracking, but the tracking mechanism tolerates a variety of flap modifications. FEN1 has a flexible loop structure through which the flap has been proposed to thread. However, efficient cleavage of branched structures is inconsistent with threading the flap through a hole in the protein.     

Models to study the pathogenesis of thyroid autoimmunity.

In vitro and in vivo models to study the pathogenesis of thyroid autoimmunity are reviewed. Animal models with experimentally induced or spontaneously developed autoimmune thyroid disease as well as transplantation models have been used extensively in these studies, but also the use of thyroid cell cultures from both humans and animals has contributed to the present state of knowledge. Cytokines may play a role in the pathogenic mechanism in thyroid autoimmunity. The major in vitro and in vivo effects of for example interleukin-1, tumour necrosis factor and gamma-interferon on differentiated thyroid cell functions are inhibitory. The advantage of using cell cultures has been the possibility of studying an influence on thyrocytes from a single agent individually, such as cytokines, hormones or growth factors. The disadvantage is that an organism is under the influence of a multitude of factors that can only be investigated in vivo in intact organisms. Both types of models have therefore been important in the understanding of thyroid autoimmunity.     

Genetic analysis of hypertropic mutants of Phycomyces

A new class of Phycomyces behavioral mutants with enhanced tropic responses has been analyzed genetically to determine the number of genes involved and the nature of their expression. These hypertropic mutants carry pleiotropic nuclear mutations. Besides their effects on sensory behavior, they also affect morphology and meiotic processes. Behavioral analyses of heterokaryons containing hypertropic and wild-type nuclei in varying proportions show that the hypertropic mutations in strains L82, L84, L86, and L88 are strongly dominant. Conversely, the hypertropic mutations carried by the strains L83, L85, and L87 are strongly recessive. We performed recombination analyses between hypertropic mutants and mutants with diminished phototropism, affected in the seven genes madA to madG. We found no evidence of linkage between the hypertropic mutations and any of these mad mutations. From crosses, we isolated double mutants carrying hypertropic mutations together with madC (night blind) and madG (stiff) mutations. The behavioral phenotypes of the double mutants are intermediate between those of the parentals. Complementation analyses show that the three recessive hypertropic mutations affect the same gene, which we call madH. The expression of the recessive hypertropic allele becomes dominant in heterokaryons carrying madC and madH nuclei; the madC gene has been implicated separately with the photoreceptor at the input to the sensory pathway, while the madH gene is associated with the growth control output. This result suggests the physical interaction of both gene products, madH and madC, in a molecular complex for the photosensory transduction chain.