Genetic Control of Mitosis: Mutation ff3Causes Premature Beginning of Telophase Chromatin Reorganization in Drosophila melanogaster

The effect of cell cycle mutation ff3 on chromosome segregation was studied on fixed cells of neural ganglia of Drosophila melanogasterlarvae. The cell distributions by diameter of interphase nuclei and by distance between sister chromatid sets were compared at anaphase and telophase. In the control wild-type strain Lausenne, the cell distribution by distance between sister chromatids in anaphase was similar to their distribution by nuclear size. The mean distance between segregating chromatids at anaphase (l _av) coincided with the mean diameter of interphase nuclei (d _av) and was 8.3 m. Cells passed to telophase when chromatids were at least 10 m apart. The mutant ff3 strain differed from the control strain Lausenne in cell distribution by interphase nuclear diameter and distance between sister chromatids in anaphase; the mean nuclear diameter and mean distance between segregating chromatids similarly increased to 9.3 m. A specific feature of mitosis in mutant strain ff3 was a premature beginning of telophase chromatin reorganization. This caused the occurrence of cells with abnormally short (less then the interphase nuclear diameter) distance between sister chromatid sets in telophase but not in anaphase, as if these cells had passed from anaphase to telophase prematurely, during the chromatid movement toward poles in anaphase A.     

The Drosophila Salivary Gland Chromocenter Contains Highly Polytenized Subdomains of Mitotic Heterochromatin

Peri-centromeric regions of Drosophila melanogaster chromosomes appear heterochromatic in mitotic cells and become greatly underrepresented in giant polytene chromosomes, where they aggregate into a central mass called the chromocenter. We used P elements inserted at sites dispersed throughout much of the mitotic heterochromatin to analyze the fate of 31 individual sites during polytenization. Analysis of DNA sequences flanking many of these elements revealed that middle repetitive or unique sequence DNAs frequently are interspersed with satellite DNAs in mitotic heterochromatin. All nine Y chromosome sites tested were underrepresented >20-fold on Southern blots of polytene DNA and were rarely or never detected by in situ hybridization to salivary gland chromosomes. In contrast, nine tested insertions in autosomal centromeric heterochromatin were represented fully in salivary gland DNA, despite the fact that at least six were located proximal to known blocks of satellite DNA. The inserted sequences formed diverse, site-specific morphologies in the chromocenter of salivary gland chromosomes, suggesting that domains dispersed at multiple sites in the centromeric heterochromatin of mitotic chromosomes contribute to polytene β-heterochromatin. We suggest that regions containing heterochromatic genes are organized into dispersed chromatin configurations that are important for their function in vivo.     

The Heterochromatic Rolled Gene of Drosophila Melanogaster Is Extensively Polytenized and Transcriptionally Active in the Salivary Gland Chromocenter

This paper reports a cytogenetic and molecular study of the structural and functional organization of the Drosophila melanogaster chromocenter. The relations between mitotic (constitutive) heterochromatin and α- and β-heterochromatin are not fully understood. In the present work, we have studied the polytenization of the rolled (rl) locus, a 100-kb genomic region that maps to the proximal heterochromatin of chromosome 2 and has been previously thought to contribute to α-heterochromatin. We show that rolled undergoes polytenization in salivary gland chromosomes to a degree comparable to that of euchromatic genes, despite its deep heterochromatic location. In contrast, both the Bari-1 sequences and the AAGAC satellite repeats, located respectively to the left and right of rl, are severely underrepresented and thus both appear to be α-heterochromatic. In addition, we found that rl is transcribed in polytene tissues. Together, the results reported here indicate that functional sequences located within the proximal constitutive heterochromatin can undergo polytenization, contributing to the formation of β-heterochromatin. The implications of this finding to chromocenter structure are discussed.     

Structural Characteristics of the Chromocenter in Ovary Cells ofc(3)Gand nodMutants of Drosophila melanogaster

Homolog pairing, chromosome morphology, and chromosome disjunction in the first meiotic division were studied in the oocytes of c(3)G/c(3)Gfemale Drosophila melanogasterat developmental stages 3–4 and 14. It was found that homologs were completely or partly paired in some cells (about 20% in either case). The lengths of chromosomes in +/+, +/c(3)G, and c(3)G/c(3)Gcells were at a ratio of 1.0 : 1.6 : 2.2. The chromocenters of homozygous cells had an abnormal structure. There was no meiotic block in metaphase 1, and chromosomes only segregated equally in about 80% of anaphases of the first meiotic division. The data obtained correspond to the abnormal variants of the formation of the chromocenter in c(3)G/c(3)Gfemales that could be predicted based on the two-ring structure of the chromocenter. The mechanism of the effect of the homo- and heterozygosity for the hypomorphic mutation c(3)Gon the formation of the synaptonemal complex (SC) and crossing over frequency was suggested. In nod/nodhomozygous females, asynapsis of pericentromeric regions of homologs was observed in the chromocenter. It was assumed that NOD kinezin is necessary at the last stages of pairing of the pericentromeric regions of homologs and formation of the coordinating bonds between them.     

Genetic Control of Development of Malpighian Tubules in Drosophila melanogaster

Malpighian tubules of insects are a functional analog of mammalian kidneys and serve as a classical model for studying the structure and functions of transport epithelium. The review contains the data on structural organization, functioning, and formation of the Malpighian tubules during embryogenesis in Drosophila melanogaster. Various systems of genes are described that control the program of development of the renal (Malpighian) tubules in D. melanogaster. A special attention is paid to the ways of signal transduction and factors involved in cell differentiation, proliferation, and morphological transformation during development of the Malpighian tubules. Evolutionarily conservative genetic systems are considered that are involved in the control of development of both the renal epithelium ofDrosophila and mammalian kidneys. A relationship was noted between the disturbed balance of genetic material and congenital defects of the human excretory system.     

Genetic and Proteomic Evidence for Roles of Drosophila SUMO in Cell Cycle Control,Ras Signaling,and Early Pattern Formation

SUMO is a protein modifier that is vital for multicellular development. Here we present the first system-wide analysis, combining multiple approaches, to correlate the sumoylated proteome (SUMO-ome) in a multicellular organism with the developmental roles of SUMO. Using mass-spectrometry-based protein identification, we found over 140 largely novel SUMO conjugates in the early Drosophila embryo. Enriched functional groups include proteins involved in Ras signaling, cell cycle, and pattern formation. In support of the functional significance of these findings, sumo germline clone embryos exhibited phenotypes indicative of defects in these same three processes. Our cell culture and immunolocalization studies further substantiate roles for SUMO in Ras signaling and cell cycle regulation. For example, we found that SUMO is required for efficient Ras-mediated MAP kinase activation upstream or at the level of Ras activation. We further found that SUMO is dynamically localized during mitosis to the condensed chromosomes, and later also to the midbody. Polo kinase, a SUMO substrate found in our screen, partially colocalizes with SUMO at both sites. These studies show that SUMO coordinates multiple regulatory processes during oogenesis and early embryogenesis. In addition, our database of sumoylated proteins provides a valuable resource for those studying the roles of SUMO in development.     

Drosophila-Host Genetic Control of Susceptibility to Drosophila C Virus

Interactions between Drosophila C virus (DCV) and its natural host, Drosophila melanogaster, were investigated using 15 geographical population samples infected by intraabdominal inoculation. These strains derived from natural populations of D. melanogaster differed in susceptibility to the DCV(C). One strain was ``partially tolerant'. Isofemale lines obtained from one susceptible and one partially tolerant strain were studied. The partially tolerant phenotype was dominant, and there was no difference between F(1) progeny of direct and reciprocal crosses. Analysis of F(2) progeny showed that neither sex-linked genes nor maternal effects are involved in susceptibility to DCV(C). The partially tolerant strain phenotype was dominant and segregated with chromosome III. Two nonexclusive hypotheses are proposed to explain chromosome III gene action.     

Genetic Control of Biogenic-Amine Systems in Drosophila Under Normal and Stress Conditions

The contents of octopamine and its precursors (tyrosine and tyramine) were studied in adults of two lines of Drosophila virilis with contrasting stress responses. It was demonstrated that in individuals responding to stress by a hormonal stress reaction (line 101), the contents of octopamine and tyrosine are lower than in nonresponding flies (line 147). It was found that there is no difference between the lines in the level of tyramine under normal conditions. The dopamine response to stressor was also studied. Genetic analysis of these differences revealed that they are controlled by a single gene and that the gene is not sex-linked. The gene controlling the response was found to be linked to chromosome 6 of D. virilis.     

Control of Female Reproduction in Drosophila: Genetic Dissection Using Gynandromorphs

The sexual behavior of Drosophila melanogaster gynandromorphs was studied to analyze the relationship between different steps in the female reproductive pathway. It was assumed that, in some gynandromorphs, certain female functions are missing because the corresponding control sites (foci) are either composed of male tissue or did not develop. A given gynandromorph can show elements of both male and female reproductive pathways. None of the steps of the female reproductive pathway appeared to be dependent on any other, in contrast to male behavior where, for example, following of females is a prerequisite for attempted copulation. By correlating each of the behaviors with the genotype of the cuticle, we confirmed previous findings that the focus for the female sex appeal is located in the abdomen, but receptivity to copulation is controlled by a site in the head. Many of the gynandromorphs did not lay eggs, presumably because either the focus controlling egg transfer from the ovaries to the uterus or the one controlling egg deposition was composed of male tissue. Many of the nonovipositing gynandromorphs laid eggs while dying or could be induced to deposit eggs after implantation of hormone-producing glands or topical application of a juvenile hormone analog. Some of the noninseminated gynandromorphs laid eggs at the rate characteristic for inseminated females, suggesting that an oviposition focus (mapping in the head region) suppresses oviposition in virgin females, but not in gynandromorphs whose focus is composed of male tissue. Some of the inseminated gynandromorphs oviposited eggs at a low rate, possibly because the focus responsible for detection of insemination could not function properly. Some of the inseminated gynandromorphs laid unfertilized eggs, revealing the importance of the focus controlling sperm release from the seminal receptacle. Foci controlling egg transfer, egg deposition and sperm release are located in the thorax, according to mosaic fate mapping results and studies on the reproductive behavior of decapitated females. The location of egg deposition in the culture vial seems to be controlled by a brain site. Sexual behavior in Drosophila does not depend on the presence (or absence) of the ovary or germ line.     

Isolation of the Chromocenter Fraction from Mouse Liver Nuclei

A new method for isolation of the constitutive heterochromatin (chromocenters) from interphase nuclei of mouse liver has been developed. This method allows separation of chromocenters of different size. Chromocenter fractions are essentially free of nucleoli and other contaminants. In contrast to nuclei and nucleoli, the chromocenter fraction is characterized by simpler protein composition, this fraction having a reduced number of proteins (especially high molecular weight proteins). Chromocenters contain all histone fractions; however, the relative proportion of histone H1 is lower and histone H3 is higher than in the total nuclear chromatin. The amount of non-histone proteins of 51, 63, 73, and 180 kD is higher in the chromocenter fraction than in nuclei and nucleoli. The use of immunocytochemistry and immunoblotting methods revealed the presence of the specific kinetochore component, CENP A protein. This suggests tight association of some molecular kinetochore components with chromocenters in the interphase.     

Genetic Control of Chromosome Elimination during Haploid Formation in Barley

Genetic control over chromosome stability in the interspecific hybrid embryos of Hordeum vulgare and H. bulbosum has been hypothesized to reside on specific chromosomes. In this study, crosses between the primary trisomic lines for the seven different H. vulgare chromosomes and tetraploid H. bulbosum revealed that both chromosomes 2 and 3 of H. vulgare were involved in the control of chromosome elimination. Subsequent crosses using the available monotelotrisomics for chromosomes 2 and 3 led to the conclusion that both arms of chromosome 2 and the short arm of chromosome 3 most likely contain major genetic factors.—From the results of this study and the genome balance observed in the interspecific crosses between H. vulgare and H. bulbosum at the diploid and tetraploid cytotypes, it appears that the factors causing the elimination of the bulbosum chromosomes are located on the H. vulgare chromosome. These factors are offset or balanced by factors on the H. bulbosum chromosomes which, when present in sufficient dosage, either neutralize the effects of the vulgare factors or are able to "protect" the bulbosum chromosomes.     

Genetic Analysis of Two Female-Sterile Loci Affecting Eggshell Integrity and Embryonic Pattern Formation in Drosophila Melanogaster

We have analyzed female-sterile mutations at the X-linked loci fs(1)Nas and fs(1)ph which show allele-specific effects on egg shell structure and embryonic pattern formation. The majority of mutant alleles at both loci lead to a collapsed egg phenotype. The maternal effect lethal phenotype is characterized by cuticle defects resembling those found in three autosomal mutants of the terminal class. We have analyzed the complementation behavior of various heteroallelic combinations at both loci and show that one such combination at the fs(1)Nas locus is capable of restoring normal fertility. We have investigated possible interactions between fs(1)Nas and fs(1)ph and also between the terminal allele of fs(1)Nas and various maternal effect mutations altering the anteroposterior polarity of embryos. We have isolated one new allele of fs(1)Nas which combines the locus-typical phenotypic features with novel cuticle phenotypes. Our results suggest that the products of fs(1)Nas and fs(1)ph are required for the stability of the vitelline membrane and are also involved in a morphogenetic pathway necessary for the correct differentiation of the terminal regions of the embryo. Possible mechanisms to account for the association of these two functions are discussed.