Human steroidogenic acute regulatory protein: functional activity in COS-1 cells, tissue-specific expression, and mapping of the structural gene to 8p11.2 and a pseudogene to chromosome 13.

Steroidogenic acute regulatory protein (StAR) appears to mediate the rapid increase in pregnenolone synthesis stimulated by tropic hormones. cDNAs encoding StAR were isolated from a human adrenal cortex library. Human StAR, coexpressed in COS-1 cells with cytochrome P450scc and adrenodoxin, increased pregnenolone synthesis > 4-fold. A major StAR transcript of 1.6 kb and less abundant transcripts of 4.4 and 7.5 kb were detected in ovary and testis. Kidney had a lower amount of the 1.6-kb message. StAR mRNA was not detected in other tissues including placenta. Treatment of granulosa cells with 8-bromo-adenosine 3',5'-cyclic monophosphate for 24 hr increased StAR mRNA 3-fold or more. The structural gene encoding StAR was mapped using somatic cell hybrid mapping panels to chromosome 8p. Fluorescence in situ hybridization placed the StAR locus in the region 8p11.2. A StAR pseudogene was mapped to chromosome 13. We conclude that StAR expression is restricted to tissues that carry out mitochondrial sterol oxidations subject to acute regulation by cAMP and that StAR mRNA levels are regulated by cAMP.

Mapping leaf surface landscapes.

Leaf surfaces provide the ecologically relevant landscapes to those organisms that encounter or colonize the leaf surface. Leaf surface topography directly affects microhabitat availability for colonizing microbes, microhabitat quality and acceptability for insects, and the efficacy of agricultural spray applications. Prior detailed mechanistic studies that examined particular fungi-plant and pollinator-plant interactions have demonstrated the importance of plant surface topography or roughness in determining the outcome of the interactions. Until now, however, it has not been possible to measure accurately the topography--i.e., the three-dimensional structure--of such leaf surfaces or to record precise changes in patterns of leaf surface elevation over time. Using contact mode atomic force microscopy, we measured three-dimensional coordinates of upper leaf surfaces of Vaccinium macrocarpon (cranberry), a perennial plant, on leaves of two age classes. We then produced topographic maps of these leaf surfaces, which revealed striking differences between age classes of leaves: old leaves have much rougher surfaces than those of young leaves. Atomic force microscope measurements were analyzed by lag (1) autocorrelation estimates of leaf surfaces by age class. We suggest that the changes in topography result from removal of epicuticular lipids and that the changes in leaf surface topography influence phylloplane ecology. Visualizing and mapping leaf surfaces permit detailed investigations into leaf surface-mediated phenomena, improving our understanding of phylloplane interactions.

A single gene for lycopene cyclase, phytoene synthase, and regulation of carotene biosynthesis in Phycomyces

Previous complementation and mapping of mutations that change the usual yellow color of the Zygomycete Phycomyces blakesleeanus to white or red led to the definition of two structural genes for carotene biosynthesis. We have cloned one of these genes, carRA, by taking advantage of its close linkage to the other, carB, responsible for phytoene dehydrogenase. The sequences of the wild type and six mutants have been established, compared with sequences in other organisms, and correlated with the mutant phenotypes. The carRA and carB coding sequences are separated by 1,381 untranslated nucleotides and are divergently transcribed. Gene carRA contains separate domains for two enzymes, lycopene cyclase and phytoene synthase, and regulates the overall activity of the pathway and its response to physical and chemical stimuli from the environment. The lycopene cyclase domain of carRA derived from a duplication of a gene from a common ancestor of fungi and Brevibacterium linens; the phytoene synthase domain is similar to the phytoene and squalene synthases of many organisms; but the regulatory functions appear to be specific to Phycomyces.

Three-dimensional cryoelectron microscopy of dimeric kinesin and ncd motor domains on microtubules.

Kinesin and ncd motor proteins are homologous in sequence yet move in opposite directions along microtubules. We have previously shown that monomeric kinesin and ncd bind in the same orientation on equivalent sites relative to the ends of tubulin sheets of known polarity. We now report cryoelectron microscope images of 16-protofilament microtubules decorated with both single- and double-headed kinesin and double-headed ncd. Three-dimensional density maps and difference maps show that, in adenosine 5'-[beta,gamma-imido]triphosphate, both dimeric motors bind tightly to microtubules via one head, leaving the other free, though apparently in a fixed position. The attached heads of dimers bind to tubulin in the same way as single kinesin heads. The second heads are connected to the tops of the first but, whereas the second kinesin head is closely associated with the first, pairs of ncd heads are splayed apart. There is also a distinct difference in orientation: the second kinesin head is tilted toward the microtubule plus end, while the second head of ncd points toward the minus end.