Mutations that perturb poly(A)-dependent maternal mRNA activation block the initiation of developmentby
|
In Drosophila, maternal effect genes are expressed during oogenesis
that produce translationally inactive mRNAs. These mRNAs are stored regionally
in the cytoplasm of the oocyte for future translation during such processes
as oocyte maturation, fertilization and early embryonic development. Some
maternal effect genes have been found to be crucial in determining embryonic
asymmetry. Drosophila mutants exist that express the same phenotypic
characteristics as those produced by selective removal of cytoplasm from
different regions of the oocyte. These mutants helped determine that the
bicoid gene is necessary for development of anterior structures such
as the head and thorax. Similarly, nanos determines posterior structures
and toll dorsoventral. These are regulatory genes producing protein
products that regulate development through transcriptional regulation of
target genes (Browder et al., 1991; Xiuguang et al., 1996).
Marshal Lieberfarb et al. (1996) studied defects in translation of
the mRNAs from these genes. They used female-sterile mutants that were defective
in translation of maternal mRNAs and found two genes, cortex and
grauzone to be crucial to both translation and polyadenylation
of bicoid and toll mRNAs. Mutants with defective cortex
genes contained bicoid mRNA identical in amount, localization and
structure as that found in wild-type embryos but with shortened poly(A)
tails. Embryos from homozygous cortex-deficient females showed profound
deficiency in Bicoid protein expression. In contrast, cortex mutants
had fully translatable nanos mRNA, because translation of this mRNA
was poly(A)-independent. Mutants with defective grauzone genes showed
results similar to cortex mutants. Therefore, the bicoid and
toll genes form a separate class of genes relying on polyadenation
for translation.
To demonstrate that polyadenation was the mechanism used in translation,
Lieberfarb et al. first showed differences in poly(A) length of wild-type
and cortex mutant mRNAs via a PCR poly(A) test. They determined that
the bicoid mRNA acquires an elongated poly(A) tail to approx. 140
nucleotides in the wild-type, whereas the mutants had their bicoid
mRNA elongated to a maximun length of approx. 80 nucleotides. This indicates
that the cortex gene is required for proper polyadenylation of cytoplasmic
bicoid mRNA. If translational activity is dependent on the length
of poly(A) tails, then injecting cortex mutants with polyadenylated
bicoid mRNA should circumvent this deficiency; Lieberbarb et al.
found that it did.
Because cortex-deficient embryos produced defects other than just
those produced by deficiency of Bicoid protein, the researchers speculated
that the gene might be required for polyadenylation and translation of a
whole class of mRNAs. The Toll protein in cortex-deficient embryos
was also reduced, so they tested whether polyadenylation oftoll mRNA
was also affected. As in the previous tests with bicoid mRNA, the
researchers found that wild-type embryos elongate toll mRNA to approx.
250 A residues, whereas cortex mutants had only about 150 poly(A)s.
This confirmed their suspicion that the cortex gene is required both
for proper polyadenylation and translation of multiple maternal mRNAs.
To demonstrate that mutants in cortex and grauzone genes did
not have a translational defect unrelated to polyadenylation, they tested
whether nanos mRNA (located in the posterior pole of the embryo and
not polyadenylated) could be translated. Their results showed that production
of the Nanos protein was the same in the mutants as in the wild-type.
The researchers also linked the cortex gene to defects in completion
of meiosis. They examined the number of nuclei and the microtubule organization
in mutant embryos and found a complete absence of microtubule organization
in mutant embryos and found a complete absence of microtubule-containing
spindles in the central regions. This indicates that initiation of mitotic
divisions was hindered. Furthermore, no polar bodies were detected in the
mutant embryos, indicating that meiosis was halted. They felt that the mechanisms
hindering these processes could well be related to polyadenylation and translation.
To determine whether a developmental defect was occurring due to somatic
mutations in embryos, rather than translational impairment of an entire
class of mRNAs, the researchers speculated that the cortex gene would
be necessary in germ cells if the latter case prevailed. In experiments
with females homozygous for a deficiency in the cortex gene of only
the germ cells, offspring had the same morphological features as demonstrated
previously in the cortex mutant phenotype. These results indicated
that a functional cortex gene is required in the germ line for polyadenylation
and mRNA translation. Temperature-sensitive mutants of females or embryos
showed that translation of this gene is required in mid- to late oogenesis
and early embryogenesis. Shifting of the females or embryos to non-permissive
temperatures at various times produced various developmental deficiencies.
The results of these studies support the conclusion that mutations in the
cortex and grauzone genes disturb the regulation of polyadenylation
of maternal mRNAs. When defective polyadenylation of maternal mRNAs results
in shortened poly(A) mRNAs, translation cannot occur, resulting in deficiencies
in oocytes of microtubule reorganization and meiosis, and deficiencies in
embryonic development such as body symmetry.
| Gametogenesis | |||||
| The Foundations of Developmental Biology | |
The Developmental Biology Journal Club | |||
Browder, L.W., Erickson, C.A and Jeffery, W.R. 1991. Developmental Biology.
Third edition. Saunders College Pub. Philadelphia (see pp. 97-98, 597-598).
Lieberfarb, M. E., Chu, T., Wreden, C., Theurkauf, W., Gergen, J.P. and
Strickland, S. 1996. Mutations that perturb poly(A)-dependent maternal mRNA
activation block the initiation of development. Development 122:
579-588.
Xiuguang, M., Yuan, D., Diepold, K., Scarborough, T. and Ma, J. 1996. The
Drosophila morphogenetic protein Bicoid binds DNA cooperatively.
Development 122: 1195-1206 (1996).
Copyright © 1996 Fabiola Aparicio, Doris Cheng, Jason Wilshusen,
and Sharon Clark.This material may be reproduced for educational purposes
only provided credit is given to the original source.
November 3, 1996