Posttranscriptional gene silencing (PTGS) is an ancient eukaryotic regulatory mechanism in which a particular RNA sequence is targeted and destroyed. The helper component-proteinase (HC-Pro) of plant potyviruses suppresses PTGS in plants. Using a yeast two-hybrid system, we identified a calmodulin-related protein (termed rgs-CaM) that interacts with HC-Pro. Here we report that rgs-CaM, like HC-Pro itself, suppresses gene silencing. Our work is the first report identifying a cellular suppressor of PTGS.
A near-fll-length casbene synthase cDNA clone, pCS7, was isolated by using a partial cDNA clone, pCS4, to probe a g10 library constructed from poly(A)+ RNA from elicited castor bean dlings. The cDNA insert had a length of 1983 bases with a polyadenylate tail of 19 bases. Translation of the cDNA sequence revealed an open reading frame encoding a 601-aa protein with a predicted Mr of 68,960. Search of the GenBank data base with the deduced translation product revealed 42% identity and 65% similarity with 5-epiaristolochene synthe from tobacco and 31% identity and 53% simarity with limonene synthase from spearmint. Each of the three proteins catalyzes an intramolecular cyclization of a prenyl diphosphate substrate to a specific cyclic terpenoid hydrocarbon product. The proposed reaction mechanisms for the three catalytic processes share common chemical features, even though the products being formed are members of three different es of terpenoid compounds. Analysis of the alignment of the three proteins suggests that both primary and secondary structural elements are conserved. These smilarties suggest that the genes that encode terpenoid cyclization enzymes of this type in anglosperms have undergone divergent evolution from an ancestral progenitor gene. In support of this proposition, the locations offive ofthe six introns in the casbene synthase gene align very closely with those of the five introns in the 5-epi-aristolochene synthase gene.Casbene is a macrocyclic diterpene hydrocarbon that serves as a phytoalexin in castor bean (Ricinus communes L.) (1). Casbene synthase (CS; EC 4.6.1.7) (previously designated casbene synthetase) from castor bean seedlings catalyzes the cyclization of geranylgeranyl diphosphate (GGPP) to form the macrocyclic hydrocarbon in a single enzymatic step. The inducible enzyme activity is found in proplastids of germinating seedlings infected by the fungus Rhizopus stolonifer (2). The plant cells recognize oligogalacturonide fragments released from pectic components of the plant cell wall by a secreted fungal endopolygalacturonase as a signal to activate CS induction (3). CS activity reaches detectable levels after 5 hr of incubation with pectic fragments, with a peak occurring after 10-12 hr (4). The active enzyme consists of a single polypeptide and has an apparent Mr of 59,000 as determined by SDS/PAGE. Polyclonal antibodies to the purified enzyme have been used to isolate a partial cDNA clone from a Agtll library (5). RNA gel blots probed with this clone show that casbene synthase mRNA is undetectable in unelicited seedlings but accumulates to a maximum at 6 hr following elicitation. Nuclear run-on experiments indicate that control is exerted at the level of transcription.A full-length CS cDNA clone was isolated as part of an effort to characterize the gene structure in order to identify cis-acting elements involved in the induction of this gene by oligogalacturonide elicitors. This report describes the characterization of this clone § and the comparison of the primary struct...
Dominant white coat colour (W) is a depigmentation syndrome, known in miscellaneous species. When homozygous in the horse (similar in mice), the mutation responsible for the white phenotype is lethal in a very early stage of gestation. It seems, that the action of the dominant white allele is not always fully penetrant, resulting occasionally in spotted look alike offspring. These horses resemble a coat colour pattern known as sabino spotting. So far, it is not known whether dominant white (W) and sabino spotting (S) share a common genetic background. In this study, a pedigree consisting of 87 horses segregating for dominant white (W) was used to genetically localize the horse (W)-locus. Microsatellite ASB23 was found linked to (W), which allowed us to map dominant white to a region on horse chromosome 3q22. Tyrosine kinase receptor (KIT) was previously mapped to this same chromosome region (3q21-22). KIT and its ligand (KITLG) are responsible for the normal function of melanogenesis, haematopoiesis and gametogenesis. So far, sequence analysis of different KIT gene fragments did not lead to new polymorphisms, except for a SNP detected in KIT intron 3 (KITSNPIn3). Additional microsatellites from ECA3q (TKY353 and LEX7), together with KITSNPIn3 allowed us to state more precisely the (W)-mutation. The positional results and comparative functional data strongly suggest that KIT encodes for the horse (W)-locus. ZusammenfassungGenetische Lokalisierung von ,,Dominant Weiss (W)ÔÔ, einem homozygot letalen Merkmal beim Pferd (Equus caballus) Die dominant weisse Fellfarbe (W) ist eine Form der Depigmentierung, die bei vielen Spezies auftritt. Beim Pferd wirkt die Mutation fü r Dominant Weiss (W) in homozygoter Form (analog zur Maus), bereits in einem sehr frü hen Stadium der Trächtigkeit letal. Es scheint, dass die Wirkung des dominant weissen Allels nicht immer mit vollständiger Penetranz erfolgt. Dies fü hrt gelegentlich zu Nachkommen mit einer Art Schecken-Fellzeichnung. Solche Pferde sind phänotypisch mit den sogenannten ,,Sabino-ScheckenÔÔ vergleichbar. Es ist bis jetzt nicht bekannt ob Dominant Weiss (W) und Sabino-Scheckung (S) einen gemeinsamen genetischen Hintergrund besitzen. Mittels eines Pedigrees aus 87 Pferden, in dem Dominant Weiss (W) segregiert, konnte in der vorliegenden Studie der equine (W)-Locus genetisch lokalisiert werden. Der Mikrosatellit ASB23 erwies sich als gekoppelt mit (W) und ermö glichte die Zuweisung des (W)-Locus auf eine Region von Chromosom ECA 3q22. Das Gen fü r den Tyrosinkinaserezeptor (KIT) liegt ebenfalls in dieser Chromosomenregion (3q21-22). Das KIT-Gen ist zusammen mit dem KIT-Liganden (KITLG) verantwortlich fü r einen normal funktionierenden Ablauf der Melanogenese, Hämatopoese und Gametogenese. Die direkte Sequenzierung von KIT-Genfragmenten fü hrte bis jetzt zu keinen neuen Polymorphismen, ausser einem SNP in KIT Intron 3 (KITSNPIn3). Mittels weiterer Mikrosatelliten von ECA3q (TKY353 and LEX7) sowie KITSNPIn3 gelang es, die (W)-Mutation genauer zu positionieren. Die vorliegend...
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