The temperature sensitivity of mutations in species of the virilis group of Drosophila. II. The effect of temperature on the maternal effect on the puffed marker following hybridization

The influence of temperature (17 and 31 degrees) on the maternal effect of mutation Puffed (Pu) in Drosophila hybrids has been studied. In the hybrids female D. littoralis +/+ x male D. virilis Pu/Pu, the stage of formation of black ring on anterior spiracles in the 3rd larval instar is sensitive to 31 degrees. In the hybrids female D. virilis Pu/Pu x male D. littoralis +/+, the expression of Pu gene manifests the maternal effect and as a result, two temperature-sensitive stages are found. The first one--onset of embryogenesis (2 to 4 hrs). At the temperature 17 degrees, the penetrance of Pu increases. The second stage is sensitive to 31 degrees and coincides with the period of black rings formation on anterior spiracles in the 3rd laval instar. It has been shown that at least two genetic systems take part in the formation of this feature. One group of genes controls the maternal effect and is sensitive to low temperature in the early embryogenesis of the hybrids female D. virilis x male D. littoralis. The second one--Pu gene and its modifiers--is active during the 2nd half of the 3rd larval instar and is heat-sensitive.     

Molecular genetic analysis of apm-2 and aps-2, genes encoding the medium and small chains of the AP-2 clathrin-associated protein complex in the nematode Caenorhabditis elegans

In this report, we analyzed the apm-2 and aps-2 genes, which encode the nematode homologues of the medium chain and the small chain of the plasma membrane-associated clathrin-associated protein complex AP-2, respectively. We determined the genomic structure of the two genes. We show that apm-2 and aps-2 genes are expressed in most, if not all, cells during embryogenesis, and that the two genes are expressed primarily in neurons and some hypodermal cells following hatching through adulthood. RNA interference experiments showed that the reduction of either apm-2 or aps-2 gene function causes embryonic lethality, larval lethality at various stages of development, and other morphological defects in the larval stages. These results indicate that apm-2 and aps-2 gene functions are required during both embryogenesis and larval stages, and that their functions may be required in proper neuronal functions.     

Characterization of New Punch Mutations: Identification of Two Additional Mutant Classes

The Punch locus of Drosophila melanogaster which encodes the pteridine biosynthetic enzyme, GTP cyclohydrolase, is genetically complex. Lethal alleles of the locus resolve into an array of interallelic complementation groups, and at least one class of mutations is developmentally specific, affecting GTP cyclohydrolase activity only in the heads of adults. All previously isolated Punch alleles were identified on the basis of a mutant eye color phenotype. By screening mutagenized chromosomes over Punch region deficiencies, we have now isolated new alleles on the basis of lethal and visible phenotypes. Most of these alleles fall into previously identified genetic classes, but two new classes of mutations were also found. One class contains two alleles that behave as dominant lethal mutations in some genetic backgrounds. The other class represents a second developmentally specific set of alleles that affect the function of the Punch locus only during embryogenesis.     

Genetic Analysis of Two Allelic Temperature-Sensitive Mutants of DROSOPHILA MELANOGASTER Both of Which Are Zygotic and Maternal-Effect Lethals

After fertilization, the development of a zygote depends upon both gene products synthesized by its maternal parent and gene products synthesized by the zygote itself. To analyze genetically the relative contributions of these two sources of gene products, several laboratories have been isolating two classes of mutants of Drosophila melanogaster: maternal-effect lethals and zygotic lethals. This report concerns the analysis of two temperature-sensitive mutants, OX736hs and PC025hs, which were isolated as alleles of a small-disc mutant, l(3)1902. These alleles are not only zygotic lethals, but also maternal-effect lethals. They have temperature-sensitive periods during larval life and during oogenesis. Mutant larvae exposed continuously to restrictive temperature have small discs. One-or two-day exposures to the restrictive temperature administered during the third larval instar lead to a homeotic transformation of the midlegs and hindlegs to the pattern characteristic of the forelegs. Mutant females exposed to the restrictive temperature during oogenesis produce eggs that can develop until gastrulation, but do not hatch.--The existence of these mutants, and one that was recently described by another group, implies that there may be a class of genes, heretofore unrecognized, whose products are synthesized during oogenesis, are essential for embryogenesis and are also synthesized during larval stages within imaginal disc cells.     

A mutation in the gene for delta-aminolevulinic acid dehydratase (ALAD) causes hypochromic anemia in the medaka, Oryzias latipes

A genetic screen for mutations affecting embryogenesis in the medaka, Oryzias latipes, identified a mutant, whiteout (who), that exhibited hypochromic anemia. The who mutant initially had the normal number of blood cells, but it then gradually decreased during the embryonic and larval stages. The blood cells in the who mutants show an elongated morphology and little hemoglobin activity. Genetic mapping localized who to the vicinity of a LG12 marker, olgc1. By utilizing the highly conserved synteny between medaka and pufferfish, we identified a gene for delta-aminolevulinic acid dehydratase (ALAD), which is the second enzyme in the heme synthetic pathway, as a candidate for who. We found a missense mutation in the alad gene that was tightly linked to the who phenotype, strongly suggesting that the hypochromic anemia phenotype in the who mutant is caused by a loss of the alad function. Thus, who mutants represent a model for the human disease ALAD-deficiency porphyria.     

Specificity of gene action during central nervous system development in Drosophila melanogaster: analysis of the lethal (1) optic ganglion reduced locus

A newly defined genetic locus designated lethal (1) optic ganglion reduced (l(1)ogre: 1-18.8, 6E1/2-6E4/5) is characterized. Four alleles have been isolated, one organismal viable and three organismal lethals. Histological analyses of these mutants at the light microscopic level have detected defects only in the developing and adult central nervous system (CNS). Examination of genetic mosaics suggests that the wild-type product of this locus may function specifically in the CNS. Analyses of staged material show that abnormalities first become apparent early in the larval period, indicating that the l(1)ogre+ gene product normally acts at or before this stage. No maternal effects were detectable. Determination of the temperature-sensitive period for lethality, of a temperature-sensitive heteroallelic combination, indicates that the l(1)ogre+ gene product also acts late in the larval period. These results show that the time of l(1)ogre+ gene action overlaps the period during which growth and assembly of the imaginal CNS occurs and are consistent with the hypothesis that l(1)ogre may act specifically in the imaginal CNS during its morphogenesis.     

L(1)trol and L(1)devl, Loci Affecting the Development of the Adult Central Nervous System in Drosophila Melanogaster

Adult optic lobes of Drosophila melanogaster are composed of neurons specific to the adult which develop postembryonically. The structure of the optic lobes and aspects of its development have been described, and a number of mutants that affect its development have been identified. The focus of every screen to date has been on disruption of adult structure or function. Although these loci were originally identified on the basis of viable mutants, some have proven capable of giving rise to lethal alleles. It seems reasonable to assume that mutants which strongly affect development of the imaginal-specific central nervous system may evidence abnormalities during the late larval or pupal stages when the adult central nervous system is undergoing final assembly and might show a lethal phase prior to eclosion (as is true for mutations at the previously defined l(1)ogre locus). We have carried out the first screen of autosomal and sex-linked late larval and pupal lethals to identify mutations that affect the development of the optic lobes. Our screen yielded nine mutants that could tentatively be grouped into three classes, depending on the neuroblast population affected and imaginal disc phenotypes. Two of these, including one that is allelic to l(1)zw1, were chosen for further analysis.     

Mutations That Alter the Timing and Pattern of Cubitus Interruptus Gene Expression in Drosophila Melanogaster

The cubitus interruptus (ci) gene is a member of the Drosophila segment polarity gene family and encodes a protein with a zinc finger domain homologous to the vertebrate Gli genes and the nematode tra-1 gene. Three classes of existing mutations in the ci locus alter the regulation of ci expression and can be used to examine ci function during development. The first class of ci mutations causes interruptions in wing veins four and five due to inappropriate expression of the ci product in the posterior compartment of imaginal discs. The second class of mutations eliminates ci protein early in embryogenesis and causes the deletion of structures that are derived from the region including and adjacent to the engrailed expressing cells. The third class of mutations eliminates ci protein later in embryogenesis and blocks the formation of the ventral naked cuticle. The loss of ci expression at these two different stages in embryonic development correlates with the subsequent elimination of wingless expression. Adults heterozygous for the unique ci(Ce) mutation have deletions between wing veins three and four. A similar wing defect is present in animals mutant for the segment polarity gene fused that encodes a putative serine/threonine kinase. In ci(Ce)/+ and fused mutants, the deletions between wing veins three and four correlate with increased ci protein levels in the anterior compartment. Thus, proper regulation of both the ci mRNA and protein appears to be critical for normal Drosophila development.     

Secondary neurons are arrested in an immature state by formation of epithelial vesicles during neurogenesis of the spider Cupiennius salei

BACKGROUND: In the spider Cupiennius salei about 30 groups of neural precursors are generated per hemi-segment during early neurogenesis. Analysis of the ventral neuromeres after invagination of the primary neural precursor groups revealed that secondary neural precursors arise during late embryogenesis that partially do not differentiate until larval stages. RESULTS: In contrast to the primary groups, the secondary invaginating cells do not detach from each other after invagination but maintain their epithelial character and form so-called epithelial vesicles. As revealed by dye labeling, secondary neural precursors within epithelial vesicles do not show any morphological features of differentiation indicating that the formation of epithelial vesicles after invagination leads to a delay in the differentiation of the corresponding neural precursors. About half of the secondary neural precursor groups do not dissociate from each other during embryogenesis indicating that they provide neural precursors for larval and adult stages. CONCLUSIONS: Secondary neural precursors are arrested in an immature state by formation of epithelial vesicles. This mechanism facilitates the production of larval neural precursors during embryogenesis. I discuss the evolutionary changes that have occured during neural precursor formation in the arthropod group and present a model for the basal mode of neurogenesis.     

Characterization of a yeast mitochondrial locus necessary for tRNA biosynthesis. Deletion mapping and restriction mapping studies

Yeast mitochondrial DNA codes for a complete set of tRNAs. Although most components necessary for the biosynthesis of mitochondrial tRNA are coded by nuclear genes, there is one genetic locus on mitochondrial DNA necessary for the synthesis of mitochondrial tRNAs other than the mitochondrial tRNA genes themselves. Characterization of mutants by deletion mapping and restriction enzyme mapping studies has provided a precise location of this yeast mitochondrial tRNA synthesis locus. Deletion mutants retaining various segments of mitochondrial DNA were examined for their ability to synthesize tRNAs from the genes they retain. A subset of these strains was also tested for the ability to provide the tRNA synthesis function in complementation tests with deletion mutants unable to synthesize mature mitochondrial tRNAs. By correlating the tRNA synthetic ability with the presence or absence of certain wild-type restriction fragments, we have confined the locus to within 780 base pairs of DNA located between the tRNAMetf gene and tRNAPro gene, at 29 units on the wild-type map. Heretofore, no genetic function or gene product had been localized in this area of the yeast mitochondrial genome.     

Loss of SEC-23 in Caenorhabditis elegans causes defects in oogenesis, morphogenesis, and extracellular matrix secretion

SEC-23 is a component of coat protein complex II (COPII)-coated vesicles involved in the endoplasmic reticulum-to-Golgi transport pathway of eukaryotes. During postembryonic life, Caenorhabditis elegans is surrounded by a collagenous exoskeleton termed the cuticle. From a screen for mutants defective in cuticle secretion, we identified and characterized a sec-23 mutant of C. elegans. By sequence homology, C. elegans has only the single sec-23 gene described herein. In addition to the cuticle secretion defect, mutants fail to complete embryonic morphogenesis. However, they progress through the earlier stages of embryogenesis, including gastrulation, and achieve substantial morphogenesis before death. We demonstrated a maternal component of SEC-23 function sufficient for progression through the earlier stages of embryogenesis and explaining the limited phenotype of the zygotic mutant. By RNA-mediated interference, we investigated the effects of perturbing COPII function during various postembryonic stages. During larval stages, major defects in cuticle synthesis and molting were observed. In the adult hermaphrodite, reduction of SEC-23 function by RNA-mediated interference caused a rapid onset of sterility, with defects in oogenesis including early maturation of the germline nuclei, probably a result of the observed loss of the GLP-1 receptor from the membrane surfaces adjacent to the developing germline nuclei.     

Genetic basis of stage-specific melanism: a putative role for a cysteine sulfinic acid decarboxylase in insect pigmentation

Melanism, the overall darkening of the body, is a widespread form of animal adaptation to particular environments, and includes bookcase examples of evolution by natural selection, such as industrial melanism in the peppered moth. The major components of the melanin biosynthesis pathway have been characterized in model insects, but little is known about the genetic basis of life-stage specific melanism such as cases described in some lepidopteran species. Here, we investigate two melanic mutations of Bicyclus anynana butterflies, called Chocolate and melanine, that exclusively affect pigmentation of the larval and adult stages, respectively. Our analysis of Mendelian segregation patterns reveals that the larval and adult melanic phenotypes are due to alleles at different, independently segregating loci. Our linkage mapping analysis excludes the pigmentation candidate gene black as the melanine locus, and implicates a gene encoding a putative pyridoxal phosphate-dependant cysteine sulfinic acid decarboxylase as the Chocolate locus. We show variation in coding sequence and in expression levels for this candidate larval melanism locus. This is the first study that suggests a biological function for this gene in insects. Our findings open up exciting opportunities to study the role of this locus in the evolution of adaptive variation in pigmentation, and the uncoupling of regulation of pigment biosynthesis across developmental stages with different ecologies and pressures on body coloration.     

A Genetic Analysis of the Pteridine Biosynthetic Enzyme, Guanosine Triphosphate Cyclohydrolase, in DROSOPHILA MELANOGASTER

Strains with mutant eye color were surveyed for levels of GTP cyclohydrolase (GTP CH), the first enzyme acting in the biosynthesis of pteridines, the pigments causing red eye color in Drosophila. Six strains were found to have reduced GTP CH activity. In five of the six strains, the reduction of activity is apparent only in the adult head of homozygous mutants. We show that mutations in Punch (2-97, Pu) have severe effects on GTP CH activity. In most cases, the reduction of activity is apparent in all tissues and stages that express the enzyme. The activity of GTP CH is shown to be closely correlated with the number of Pu+ genes in the genome. One ethyl methanesulfonate (EMS)-induced Pu mutant has a GTP CH enzyme that is unstable when compared with the wild-type enzyme. Mutations in Pu fall into three general classes. The largest class has a recessive lethal and eye color phenotype, 50% or higher GTP CH activity in heterozygotes, and equivalent defects in all tissues. A second class is dominant in eye color phenotype and recessive lethal, with less than 50% GTP CH activity in heterozygotes. The third class is homozygous viable and has severe reduction of activity in the adult head, but no or less severe loss in other tissues.     

Projections of Drosophila multidendritic neurons in the central nervous system: links with peripheral dendrite morphology

Neurons establish diverse dendritic morphologies during development, and a major challenge is to understand how these distinct developmental programs might relate to, and influence, neuronal function. Drosophila dendritic arborization (da) sensory neurons display class-specific dendritic morphology with extensive coverage of the body wall. To begin to build a basis for linking dendrite structure and function in this genetic system, we analyzed da neuron axon projections in embryonic and larval stages. We found that multiple parameters of axon morphology, including dorsoventral position, midline crossing and collateral branching, correlate with dendritic morphological class. We have identified a class-specific medial-lateral layering of axons in the central nervous system formed during embryonic development, which could allow different classes of da neurons to develop differential connectivity to second-order neurons. We have examined the effect of Robo family members on class-specific axon lamination, and have also taken a forward genetic approach to identify new genes involved in axon and dendrite development. For the latter, we screened the third chromosome at high resolution in vivo for mutations that affect class IV da neuron morphology. Several known loci, as well as putative novel mutations, were identified that contribute to sensory dendrite and/or axon patterning. This collection of mutants, together with anatomical data on dendrites and axons, should begin to permit studies of dendrite diversity in a combined developmental and functional context, and also provide a foundation for understanding shared and distinct mechanisms that control axon and dendrite morphology.     

A Functional and Structural Analysis of the Sex Combs Reduced Locus of Drosophila Melanogaster

We have undertaken a developmental genetic analysis of the homeotic gene Sex combs reduced (Scr) of Drosophila melanogaster by examining embryonic and adult phenotypes of mutations affecting Scr gene function. Molecular mapping of Scr breakpoint lesions has defined a segment of greater than 70 kb of DNA necessary for proper Scr gene function. This region is split by the fushi tarazu (ftz) gene, with lesions affecting embryonic Scr function molecularly mapping to the region proximal (5') to ftz and those exhibiting polyphasic semilethality predominantly mapping distal (3') to ftz. Gain-of-function mutations are associated with genomic rearrangements and map throughout the Scr locus. Our analysis has revealed that the Scr locus encompasses genetic elements that are responsible for functions in both the embryonic and larval to adult periods of development. From these studies, we conclude that Scr is a complex genetic locus with an extensive regulatory region that directs functions required for normal head and thoracic development in both the embryo and the adult and that the regulation of Scr during these two periods is distinct.