How do single homeotic genes control multiple segment identities?

Each of the homeotic genes of the bithorax complex of Drosophila defines the identities of more than one body segment. The mechanisms by which this occurs have been elusive. In a recent report, Castelli-Gair and Akam⁽¹⁾ analyze in detail the control of parasegment 5 and parasegment 6 identities by the bithorax complex gene Ubx. Their results indicate that differences in the spatial and temporal expression patterns of Ubx are critical in determining differences between these parasegments. However, dose effects observed by others indicate that parasegment-specific differences in the level of Ubx expression are also important. For the other BX-C genes, parasegment-specific expression of protein isoforms, or combinatorial control dependent on the expression patterns of other spatially restricted regulators, may also play a role.     

Maternal expression of genes that regulate the bithorax complex of Drosophila melanogaster

A relatively large number of genes have been described that are required for the normal spatial expression of the genes of the bithorax complex. Most of these regulators appear to act negatively and are required to prevent indiscriminate expression of bithorax complex (BX-C) functions. In this report we examine five negative BX-C regulators to determine whether these are maternally expressed in germ-line derived cells. The genes studied include Additional sex combs (Asx), Polycomblike (Pcl), Sex comb extra (Sce), Sex comb on midleg (Scm), and lethal(4)29 [l(4)29]. The maternal germ-line dependent expression of each of these genes is assessed by comparison of zygotes from mothers whose functional germ cells carry no wild-type alleles to zygotes from mothers whose germ cells contain one wild-type allele. Because mutant alleles of each of the genes studied are recessive lethals, mosaic females with homozygous or hemizygous mutant germ lines were produced by pole cell transplantation. The results demonstrate that all of the negative regulators tested are expressed in the maternal germ line and all play important roles in the regulation of BX-C activities during embryogenesis. The absence of maternally supplied products from all of the genes studied except l(4)29 can be largely or completely compensated for by the activity in the zygote of a paternally contributed wild-type allele. It is argued that, with the exception of l(4)29, the genes studied in this report are qualitatively similar in function to the previously described BX-C regulators Pc, esc, and sxc. The available evidence indicates that genes within this group have functions that are not restricted to the regulation of genes that control segmental identity.     

Effects of Polycomb Group Mutations on the Expression of Ultrabithorax in the Drosophila Visceral Mesoderm

The Polycomb group (PcG) genes encode repressors of many developmental regulatory genes including homeotic genes and are known to act by modifying chromatin structure through complex formation. We describe how Ultrabithorax (Ubx) expression is affected by the PcG mutants in the visceral mesoderm. Mutant embryos of the genes extra sex combs (esc), Polycomb (Pc), additional sex combs (Asx) and pleiohomeotic (pho) were examined. In each mutation, Ubx was ectopically expressed outside of their normal domains along the anterior-posterior axis in the visceral mesoderm, which is consistent with the effect of PcG proteins repressing the homeotic genes in other tissues. All of these four PcG mutations exhibit complete or partial lack of midgut constriction. However, two thirds of esc mutant embryos did not show Ubx expression in parasegment 7 (PS7). Even in the embryos showing ectopic Ubx expression, the level of Ubx expression in the PcG mutations was weaker than that in normal embryos. We suggest that in PcG mutations the ectopic Ubx expression is caused by lack of PcG repressor proteins, while the weaker or lack of Ubx expression is due to the repression of Ubx by Abd-B protein which is ectopically expressed in PcG mutations as well.     

Maternal and zygotic requirement for thepolyhomeotic complex genetic locus inDrosophila

Summary The complex genetic locuspolyhomeotic (ph) is a member of thePolycomb (Pc)-group of genes and as such is required for the normal expression of ANT-C and BX-C genes. It also has probably other functions since amorphicph alleles display a cell death phenotype in the ventral epidermis of 12-h-old embryos. Here it is shown that lethal alleles ofph (amorph and strong hypomorph) show transformation of most of their segments towards AB8. Theph ⁺ product is required autonomously in imaginal cells. The total lack ofph ⁺ function prevents viability of the cuticular derivatives of these cells.ph has a strong maternal effect on segmental identity and epidermal development that can not be rescued by one paternally supplied dose ofph ⁺ in the zygote. These phenotypes differ substantially from those of previously describedPc-group genes. AmongPc-group genes,ph seems to be the only one that is strongly required both maternally and zygotically for normal embryonic development.     

Specificity of embryonic lethal mutations in Drosophila analyzed in germ line clones

Summary Only a small fraction of the known mutations causing death to homozygous Drosophila produce gross morphological defects during embryogenesis. We have examined fourteen such loci on the X-chromosome to determine: 1) whether the requirement for their respective activities is restricted to embryogenesis; and 2) whether the embryonic phenotype in mutant embryos is affected by the dosage of wild-type alleles in the mother. For two alleles per locus germ line clones were produced during larval development by irradiating females heterozygous for the lethal mutation and a dominant female sterile (ovo^D). Only one of the 14 loci (armadillo) is required during development of the germ cell to make morphologically normal eggs. Mutations at two other loci, (bazooka and Notch), allow normal oogenesis but cause major reductions in the viability of genetically normal (i.e., heterozygous) progeny. The majority of the loci (11/14) are not required in the germ line for either oogenesis or embryogenesis. However, in three cases (extradenticle, faintoid and lethal myospheroid), germ line homozygosity results in a readily detectible enhancement of embryonic phenotype over that observed in embryos derived from heterozygous mothers still possessing one wild type allele. The same six loci which show the most substantial effects on germ line homozygosity (arm, baz, N, exd, ftd and mys) also show an amelioration of the mutant phenotypes when maternal dosage is increased to wild type levels by using attached-X females. Four of these same loci (arm, baz, N and exd were cell lethal in imaginal discs.     

Epigenotype switching of imprintable loci in embryonic germ cells

 Expression of imprinted genes is dependent on their parental origin. This is reflected in the heritable differential methylation of parental alleles. The gametic imprints are however reversible as they do not endure for more than one generation. To investigate if the epigenetic changes in male and female germ line are similar or not, we derived embryonic germ (EG) cells from primordial germ cells (PGCs) of day 11.5 and 12.5 male and female embryos. The results demonstrate that they have an equivalent epigenotype. First, chimeras made with EG cells derived from both male and female embryos showed comparable fetal overgrowth and skeletal abnormalities, which are similar to but less severe than those induced by androgenetic embryonic stem (ES) cells. Thus, EG cells derived from female embryos resemble androgenetic ES cells more than parthenogenetic cells. Furthermore, the methylation status of both alleles of a number of loci in EG cells was similar to that of the paternal allele in normal somatic cells. Hence, both alleles of Igf2r region 2, Peg1/Mest, Peg3, Nnat were consistently unmethylated in EG cells as well as in the primary embryonic fibroblasts (PEFs) rescued from chimeras. More strikingly, both alleles of p57kip2 that were also unmethylated in EG cells, underwent de novo methylation in PEFs to resemble a paternal allele in somatic cells. The exceptions were the H19 and Igf2 genes that retained the methylation pattern in PEFs as seen in normal somatic tissues. These studies suggest that the initial epigenetic changes in germ cells of male and female embryos are similar. Received: 1 September 1997 / Accepted: 15 October 1997     

The maternal and zygotic roles of the gene Polycomb in embryonic determination in Drosophila melanogaster

The mutation Polycomb (Pc) is known to cause a variety of intersegmental transformations in homozygous and heterozygous individuals of Drosophila melanogaster; Pc⁺ is thought to act as a negative regulator of genes of the bithorax complex. The function of this gene in the maternal germ line has been assessed by examining the variation in expression of these homoeotic phenotypes in individuals derived from a maternal germ line with a single or no dose of the Pc⁺ allele. Mosaic individuals with a homozygous or heterozygous Pc germ line were produced by transplantation of pole cells, the embryonic precursors of the germ line. By employing an X-linked dominant female-sterile mutation, the identification of mosaic females and the study of progeny derived from the exogenous germ line were greatly simplified; the advantages of this system for the transplantation of pole cells for such analyses are described. In general, all thoracic and abdominal segments of homozygous Pc embryos differentiate characteristics of the eighth, most posterior, abdominal segment. The extent and uniformity of this transformation as well as other manifestations of the homozygous Pc genotype are described and shown to be correlated with the maternal germ line genotype; homozygous Pc embryos derived from a homozygous Pc maternal germ line show greater expression of these phenotypes than do genetically identical embryos derived from a heterozygous Pc maternal germ line. The expression of some homoeotic phenotypes typical of heterozygous Pc adults shows only a slight correlation with the maternal genotype, while no homoeotic transformations are clearly evident in heterozygous larvae of either origin. Thus, the maternal effect of Pc is rescuable. The results suggest that the Pc⁺ gene is active in the maternal germ line but that the absence of the maternally derived Pc⁺ product can be largely compensated by the introduction of a wild-type allele upon fertilization; this rescue indicates that the maternal activity of Pc⁺ plays no major role in the normal process of embryonic segmental determination. The normal fertility of males and females with a homozygous Pc germ line and of their progeny suggests that Pc⁺ plays no role in the determination or development of the germ line in either the maternal or zygotic genome.     

Characterization of a region of the X chromosome of Drosophila including multi sex combs (mxc), a Polycomb group gene which also functions as a tumour suppressor

Genetic analysis of the 8D3;8D8-9 segment of the Drosophila melanogaster X chromosome has assigned seven complementation groups to this region, three of which are new. A Polycomb group (Pc-G) gene, multi sex combs (mxc), is characterized and mutant alleles are described. Besides common homeotic transformations characteristic of Pc-G mutants that mimic the ectopic gain of function of BX-C and ANT-C genes, mxc mutants show other phenotypes: they zygotically mimic, in males and females, the characteristic lack of germ line seen in progeny of some maternal effect mutants of the so-called posterior group (the grandchildless phenotype). Loss of normal mxc function can promote uncontrolled malignant growth which indicates a possible relationship between Pc-G genes and tumour suppressor genes. We propose that gain-of-function of genes normally repressed by the wild-type mxc product could, in mxc mutants, give rise to an incoherent signal which would be devoid of meaning in normal development. Such a signal could divert somatic and germ line developmental pathways, provoke the loss of cell affinities, but allow or promote growth.     

Maternal effects of zygotic mutants affecting early neurogenesis inDrosophila

Summary The size of the neurogenic region ofDrosophila melanogaster is under the control of several genes of zygotic expression. Lack of function from any of those genes produces an increase of the size of the neurogenic region at the expense of the epidermal anlage. However, differences exist in the extent of neuralisation achieved by each of the genetic loci upon mutation. The present results show that in the case ofN andmam phenotype differences are due to different contributions of maternal gene expression. This could be shown by studying the phenotype which appeared in mutant embryos when the oocytes developed from homozygous mutant precursor cells. Clones of mutant cells were induced in the germ line of females heterozygous for the neurogenic mutationin trans over germ line dependent, dominant female sterile mutations. After removing maternal information the phenotype ofN andmam mutants became identical in both cases. Furthermore maternal information fromN ⁺ was found to be necessary for viability of the wildtype.     

Two Drosophila learning mutants, dunce and rutabaga, provide evidence of a maternal role for cAMP on embryogenesis

The dunce gene of Drosophila melanogaster encodes a cAMP-specific phosphodiesterase (form II). Mutant dunce flies have elevated levels of cAMP and exhibit a number of defects including learning deficiencies and female sterility. Two partial suppressors of the female sterility phenotype have been selected in an X chromosome containing a dunce null mutation. Both suppressors are associated with reduced AC2 activity. Complementation analyses suggest that both are alleles of the learning mutant rutabaga. Females homozygous for dunce null mutations that abolish PDE activity do not deposit eggs. The suppressors exhibit differential effects on egg deposition and production of progeny; double-mutant females deposit many eggs that fail to hatch, but some develop to adults. These adult progeny exhibit morphological defects that are confined mostly to the second and third thoracic segments or to the first five abdominal segments. These observations demonstrate that the dunce gene is required in adult females for egg laying and that the dunce gene provides an essential maternal function required for normal development of the zygote. Clonal analysis, employing the dominant female-sterile mutation ovoD1, demonstrates that the former requirement for PDE activity resides in somatic cells and that the latter requirement resides in germ line cells. Female germ line cells homozygous for a dunce null mutation produce oocytes that fail to develop. Thus, homozygous dunce null-mutant zygotes develop to adults solely because of the enzyme or mRNA present in the oocytes of heterozygous mothers. Mutant alleles of rutabaga act in the germ line cells to partially suppress the developmental defects caused by dunce mutations. Thus the rutabaga gene, as well as the dunce gene, functions in both somatic and germ line cells.     

hunchback, a gene required for segmentation of an anterior and posterior region of the Drosophila embryo

The locus hunchback (hb) is a member of the gap class of segmentation genes of Drosophila. A number of X-ray-induced deletions locate the hb locus at the chromosomal site 85A3-B1, to the right of the pink locus, which maps in the same interval. A total of 14 EMS and 3 X-ray-induced hb alleles have been studied. Homozygous mutant embryos show deletions of segments in two separate regions. In the six strong alleles, the labium and all three thoracic segments are deleted anteriorly while posteriorly the 8th abdominal segment and adjacent parts of the 7th abdominal segment are lacking. The eight weak alleles show smaller deletions both in the thoracic and posterior abdominal region. In the weakest allele only part of the mesothorax is deleted. Three hb alleles produce a homoeotic transformation: superimposed on a strong or weak deletion phenotype, head or thoracic segments are transformed into abdominal segments, respectively. This suggests that hb might also be involved in the regulation of genes in the Bithorax complex (BX-C). Fate mapping of the normal-appearing segments in strong mutant embryos using the UV-laser beam ablation technique (Lohs-Schardin et al., 1979) shows that these segments arise from the normal blastoderm regions. The mutant phenotype can be recognized soon after the onset of gastrulation in a failure to fully extend the germ band. In 6-hr-old mutant embryos, two clusters of dead cells are observed in the thoracic and posterior abdominal region. These observations indicate region specific requirement of hb gene function. The analysis of germ line chimeras by transplantation of homozygous mutant pole cells shows that hb is already expressed during oogenesis. Homozygous mutant embryos derived from a homozygous mutant germ line have a novel phenotype. The anterior affected region is enlarged, including all three gnathal segments and the anterior three abdominal segments. In addition three abdominal segments with reversed polarity are formed between the remaining head structures and the posterior abdomen. Heterozygous mutant embryos derived from a homozygous mutant germ line develop normally, indicating that maternal gene expression is not required for normal development.     

The multi sex combs gene of Drosophila melanogaster is required for proliferation of the germline

We present a genetic analysis showing that the Drosophila melanogaster gene multi sex combs (mxc; Santamaria and Randsholt 1995) is needed for proliferation of the germline. Fertility is the feature most easily affected by weak hypomorphic mutations of this very pleiotropic locus. Pole cell formation and early steps of gonadogenesis conform to the wild-type in embryos devoid of zygotic mxc ⁺ product. mxc mutant gonad phenotypes and homozygous mxc germline clones suggest a role for mxc ⁺ in control of germ cell proliferation during the larval stages. mxc ⁺ requirement is germ cell autonomous and specific in females, whilst in males mxc ⁺ product is also needed in somatic cells of the gonads. Although mxc can be classified among the Polycomb group (Pc-G) of genes, negative trans-regulators of the ANT-C and BX-C gene complexes, germline requirement for mxc appears independent of a need for other Pc-C gene products, and mxc gonad phenotypes are different from those induced by mutations in BX-C genes. We discuss the possible functions of the mxc ⁺ product which helps to maintain homeotic genes repressed and prevents premature larval haemocyte differentiation and neoplasic overgrowth, but promotes growth and differentiation of male and female gonads.F.D. and O.S. should be considered as equal first authors     

Preblastoderm mosaics of mutants of the bithorax-complex

Summary InDrosophila melanogaster, segmental specification takes place in groups of cells around the blastoderm stage. This segmental specification requires the function of the genes of the bithorax-complex. We have studied preblastoderm mosaics (gynandromorphs) of mutant (bx ³,pbx, Ubx, Ubx ⁸⁰) and wildtype (heterozygotes for these alleles) cells. The results show a total cell autonomy in the differentiation of both wildtype and homoeotially transformed cells. However, several unexpected phenotypes were found. They are discussed in terms of the function of the bithorax genes and early interactions between mutant and wildtype territories.     

Recessive lethal mutations within the bithorax-complex in Drosophila

Summary Genetic deficiencies of the bithorax-complex (BX-C) in Drosophila, have been used to recover recessive lethal mutations in this chromosome region following mutagenesis. Complementation analysis separates these lethal mutations into five groups within a smaller deficiency, though to remove the entire BX-C, and into 20 to the left and 4 to the right of the region. Homozygotes for each of only three groups of lethals, Ubx, abdA and AbdB, produce homoeotic segmental transformations in embryos. The functional domains of abdA and AbdB have been defined by changes in the appearance of larval hypodermal structures and of clones in imaginal tissue. The function abdA is required in all the compartments caudal to the anteroposterior border of abdominal segment 1 up to and including the anterior region of abdominal segment 8, whilst AbdB is required in abdominal segments 5 to 9. One allele of AbdB produces a ninth abdominal setal band and structures characteristic of head segments posterior to A8. Rare adult survivors hemizygous for an AbdB allele have eight abdominal segments in both sexes, and lack genitalia in females. Our findings are discussed in the context of the organisation of genetic functions within the BX-C.     

Interactions of thePolycomb group of genes with homeotic loci ofDrosophila

Summary Members of thePolycomb (Pc) group of genes are required for the correct determination of segment identity, and are thought to be negative regulators of thebithorax andAntennapedia complexes. This hypothesis has been tested molecularly for only some members of thePc group. Here, we examine the distribution ofUltrabithorax (Ubx),Antennapedia (Antp), andSex combs reduced (Scr) proteins in the epidermis, central nervous system, and midgut of embryos homozygous for mutations in tenPc group genes. We show that zygotic loss of mostPc group genes causes ectopic expression ofUbx andAntp, but that there are differences in time and tissue-specificity. FivePc group mutations lack midgut constrictions and also exhibit ectopic or suppressedUbx expression and suppression ofAntp expression. Distribution ofAntp is upset earlier than distribution ofUbx in the central nervous system of everyPc group mutant affecting both genes. Loss of the zygotic products ofPolycomb, extra sex combs, andAdditional sex combs cause ectopic expression ofScr in epidermis, and allPc group genes exceptPsc have suppressedScr expression in the nervous system. These results are discussed with respect to the function of thePc group.     

Genetic and molecular analysis of fs(1)h, a maternal effect homeotic gene in Drosophila

Mutations at the Drosophila melanogaster locus female sterile (1) homeotic (fs(1)h) result in segmental abnormalities including missing organs and homeotic transformations in the progeny of mutant mothers. Homeotic transformations are enhanced when the zygotes carry one of several third chromosome mutations, specifically alleles or deficiencies of the trithorax (trx) locus, also called Regulator-of-bithorax, and some alleles of bithorax complex (BX-C) genes. These observations suggest that maternally derived fs(1)h+ product is required, in interaction with trx and BX-C genes, for normal segment specification. The fs(1)h gene and an adjacent gene, lethal (1) myospheroid (l(1)mys), have been cloned by chromosomal walking. Mutations of fs(1)h were found within a 13-kb stretch of DNA. Poly(A)+ RNAs migrating as a doublet at 7.6 kb and a single band at 5.9 kb, which are homologous to the fs(1)h+ chromosomal region, are found in ovaries and early embryos. The largest RNAs are derived from a 20-kb chromosomal region encompassing the sites of all mapped fs(1)h alleles.     

More mog genes that influence the switch from spermatogenesis to oogenesis in the hermaphrodite germ line of Caenorhabditis elegans

The Caenorhobditis elegans XX animal possesses a hermaphrodite germ line, producing first sperm, then oocytes. In this paper, we report the genetic identification of five genes, mog-2, mog-3, mog-4, mog-5, and mog-6, that influence the hermaphrodite switch from sper-matogenesis to oogenesis. In mcg-2-mog-6 mutants, spermatogenesis continues past the time at which hermaphrodites normally switch into oogenesis and no oocytes are observed. Therefore, in these mutants, germ cells are transformed from a female fate (oocyte) to a male fate (sperm). The fem-3 gene is one of five genes that acts at the end of the germline sex determination pathway to direct spermatogenesis. Analyses of mog;fem-3 double mutants suggest that the mog-2-mog-6 genes act before fem-3; thus these genes may be in a position to negatively regulate fem-3 or one of the other terminal regulators of germline sex determination. Double mutants of fem-3 and any one of the mog mutations make oocytes. Using these double mutants, we show that oocytes from any mog;fem-3 double mutant are defective in their ability to support embryogenesis. This maternal effect lethality indicates that each of the mog genes is required for embryogenesis. The two defects in mog-2-mog-6 mutants are similar to those of mog-1: all six mog genes eliminate the sperm/oocyte switch in hermaphrodites and cause maternal effect lethality. We propose that the mog-2-mog-6 mutations identify genes that act with mog-1 to effect the sperm/oocyte switch. We further speculate that the mog-1-mog-6 mutations all interfere with translational controls of fem-3 and other maternal mRNAs. © 1993 Wiley-Liss, Inc.