Up to 36% of Neurospora crassa transformants showing an albino phenotype were recovered by transforming a wild-type strain with different portions of the carotenogenic albino-3 (al-3) and albino-1 (al-1) genes. The presence of the exogenous sequences (which were randomly integrated in ectopic locations) provoked a severe impairment in the expression of the endogenous al-1 or al-3 genes. This phenomenon, which we have termed 'quelling', was found to be spontaneously and progressively reversible, leading to wild-type or intermediate phenotypes. The phenotypic reversion is characterized by a progressive release of the transcriptional inhibition and seems to correlate with a reduction of the number of the ectopic integrated sequences. Moreover, quelling appears to be monodirectional, as, once relieved, it cannot take place again, despite the continuing presence of some of the ectopic sequences in the genome.

In barley (Hordeum vulgare L.) the unit of inflorescence is the spikelet, which bears a fertile bract, the lemma, and the floret consisting of palea, two lodicules, three stamens and the pistil. The Hooded mutation causes the appearance of an extra flower of inverse polarity on the lemma. This phenotype is governed by the single dominant genetic locus K3. Here we show that the homeobox gene Knox3 represents this locus. Ectopic Knox3 gene expression in the primordium of the extra floret is caused by a 305-base pair duplication in intron 4, and phenocopies of the mutation are obtained in the heterologous tobacco system by Knox3 overexpression. It is concluded that homeotic genes of the Knox gene family are involved in floral evocation. Furthermore, the study of polarity of reproductive organs in K and related mutants can now focus on homeobox genes.

The albino-3 (al-3) gene of Neurospora crassa, which probably encodes the carotenoid biosynthetic enzyme geranylgeranyl pyrophosphate synthetase, was cloned. The N. crassa triple mutant al-3 qa-2 aro-9 was transformed to qa-2+ with mixtures of plasmids bearing N. crassa DNA inserts, and the transformants were screened for the al-3+ phenotype. One al-3+ qa-2+ transformant (AL3-1) was examined in detail and shown to contain intact vector sequences integrated into the N. crassa genome. The vector and some flanking sequences were recovered from AL3-1 after restriction, ligation, and selection of chloramphenicol-resistant transformants of Escherichia coli. The flanking sequences were subsequently used to detect the al-3-containing plasmid in the mixture of about 1,800 plasmids. Restriction fragment length polymorphism mapping was carried out to confirm the identity of the cloned fragment. The level of the al-3 mRNA was shown to be increased 15-fold in light-induced (compared with that in dark-grown) wild-type mycelia. The light-dependent increase in al-3 mRNA levels was not observed in presumed regulatory mutant (white collar) strains.

When a wild-type strain of Neurospora crassa is transformed with different portions of the carotenogenic albino 1 or albino 3 genes, up to 30-35% of the transformants show an albino phenotype. The albino transformants presented a variety of phenotypes ranging from white or yellow to dark yellow colour. The ectopically integrated sequences provoke a severe impairment of the expression of the endogenous al-1 or al-3 genes. This phenomenon, that has been termed quelling, is found to be spontaneously and progressively reversible. In fact, all of the albino transformants have an unstable phenotype and revert progressively to wild type or intermediate phenotypes over a prolonged culturing time. The phenotypic reversion is characterised by a progressive release of the transcriptional inhibition and seems to correlate with the reduction of the number of the ectopic integrated sequences. However, there is no strict correlation between the copy number of the ectopic sequences and the intensity of quelling, as indicated by the existence of albino transformants containing only 1-2 ectopic sequences. The nature of the molecular events determining the onset of quelling is unclear, in any event, these are likely to involve some kind of interaction between the resident genes and ectopically integrated exogenous sequences. Recent evidences on a possible mechanism are presented.

No abstract