The Role of DCL1 (=DICER-LIKE1) – a Complex Gene in Arabidopsis Development

Each trait among myriad is controlled by a single or multiple genes. As soon as mutation occurs with respect to a gene, the trait (phenotype), it controls changes. The phenotypic change may be visible or invisible at the first filial generation, depending upon, whether the mutation is dominant or recessive. In case it is recessive, the expression of the trait is deferred until the following generation i.e., F2; those F2 individuals, where only recessive genes are present, display the recessive-gene-controlled trait.

Since a gene is composed of a large number of nucleotides, mutations may affect base pairs of the same- or different sites of the same gene. When mutations affect different sites of a gene, phenotypes may be unrecognizably different from the original mutant trait. For instance, the mutated chlorophyll gene, expresses itself in several shades of green pigmentation, the extreme form being an albino plant. Leaves of mutants could be half-albino, viriscent , or their blades showing alternate green and white stripes etc. However, since the color is leaf-related, it is not difficult to infer that probably these seemingly different traits are controlled by different alleles of the same gene.

The situation becomes particularly complex, when the gene of interest plays a wide-ranging regulatory role and mutations of such genes exert pleiotropic effect, as demonstrated by the multidomain RNA-helicase/nuclease gene DCL1. Earlier, it was believed, that mutations such as (a) defective embryos (embryo defective = emb76) , (b) lack of growth of outer integuments, exposing the inner integument (short integument = sin1), (c) abnormal suspensor cells development (suspensor1 – sus1) and (d) the continuous growth of the meristem without further differentiation (carpel factory =CAF1) ) arose as a result of mutations, occurring at different loci. Results of recent studies show that all the diverse phenotypes, enlisted above, arose as a result of mutations, involving the same locus (gene).

In a review article published in the November, 2002 issue of Trends in Plant Science (Vol.7: 487-91) Animesh Ray at Keck Graduate Institute, CA, and his associates at three different institutes (OSU; MCDB, UCLA; and Rochester university), have renamed this gene, DICER-LIKE1 (DCL1) on the basis of structural similarity of the predicted protein encoded by this gene with the DICER protein of Drosophila melanogaster and DCR-1 of Caenorhabditis elegans. DCR of Drosophila and DCR-1 of Caenorhabditis have been shown to encode a complex RNA-processing enzyme.

sin1-2 – a mutant allele involving short integument (sin1): Study of this allele revealed that it regulates the pattern of embryo development. Embryo development was found to be entirely dependent upon the genotype of the maternal diploid tissue surrounding the embryo. The genotype of the pollen parent, carrying the dominant allele, did not have any effect on the phenotype of the embryo. Leaves in the heterozygotes, sin1-2/SIN1 into the Columbia background were normal; so also their normal looking ovules. The latter developed into seeds but lacked viable embryos, confirming that their full cycle of development is determined by the genotype of the nucellus, surrounding the developing embryo.

These results led the authors  suggest that DCL1 plays an essential role in diploid maternal cells, orchestrating various steps involved in normal embryogenesis, and that the normal allele in the pollen carrying the dominant allele cannot compensate for this deficiency. The authors cited a few more instances from Arabidopsis to prove the essentiality of DCL1 on the determination of meristem fate i.e., signaling the transition of a vegetative- to a floral meristem.

Characterization of the DCL1 gene: The DCL1 gene, encoding for 1909 amino acids, was identified and cloned with the help of a T-DNA insetion in the caf-1 allele. A nuclear localization signal (NLS), and a second double stranded RNA  (dsRNA)-binding domain at the C-terminus distinguishes plant DCL1 from other DCR proteins of animal origin. Aided by computer searches, the authors have described six plant Dicer gene family members, four in Arabidopsis and two in rice. The same number (6) has been reported in six non-plant Dicer gene family.

Characteristics of Dicer protein: The Drosophila Dicer protein cleaves large dsRNA into fragments of 21-25. These small stretches of RNAs, are called short interfering RNAs, abbreviated to siRNAs. They target homologous RNAs and by destroying them, they bring about post-transcriptional gene-silencing. Dicer also cleaves small i.e., about 100 bp long non-coding dsRNA hairpins into 21-25 nucleotide long single-stranded RNA fragments.

Model of DICER-LIKE1 function in plants: Based on their own experience and on findings reported earlier, the authors propose an essentially important role of a small RNA- mediated pathway involving DCL1. The pathway is operational during plant development involving regulation of a range of important genes. Interestingly, in Arabidopsis, double mutants, disrupted in two closely related ARGONAUTE gene-family members (argonaute1 and pinhead/zwille), show phenotypes of dcl1 null mutants (= genes not expressed). In the double mutant, cells proliferate without showing any differentiation. In addition, weak dcl1 loss-of-function mutants and argonaute1 or pinhead/zwille single mutants were found to share phenotypic similarities such as thin sepals and petals, antherless stamens, tube- shaped leaves, loss of axillary meristems, and filaments on the stems.

Small RNAs as developmental signals: Recent studies indicate that small RNAs play a significant role in plant development as demonstrated by developmental abnormalities associated with dcl1, argonaute1 and pinhead/zwille mutants. Another important observation made in this connection is that DCL1 can function by sending its signals from a distance i.e., from the nucellus (= maternal tissue) into the developing embryo. It is believed that DCL1 signals travel through the plasmodesmata to the site, where it regulates different stages of embryo development. It is believed that the RNA-silencing mechanism creates siRNAs and the latter, in turn silences homologous RNA. Furthermore, functioning of small RNAs is probably dependent upon a DCL1 signaling, indicating that RNAs or their precursors cannot work by themselves i.e., non-autonomously. They require DCL1 for their activation for transformation of the vegetative shoot into a floral meristem.

Summing up, the authors propose that (a) a small RNA-mediated pathway involving DCL1, operates during plant development to regulate a range of important genes and (b) genes, previously designated emb76, sin1, sus1, caf1, control floral transition and floral development and they represent the mutant alleles of one single gene, namely, DCL1. The authors conclude that in spite of a number of recent exciting findings on the elucidation of the DCL1 locus and its combinatorial role with small RNAs, there is still adequate scope for further intensive research to discover mutant alleles involving this locus. Discovery of such alleles may shed further light as to how this locus functions in conjunction with small RNAs to initiate transforma- tion of vegetative- into floral meristems and promote proper growth and development of reproductive organs  such as stopping unregulated growth of different floral parts etc.

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