Eliezer Lifschytz

Gene expression and the control of spermatid morphogenesis in Drosophila melanogaster

Abstract The genetic control of spermatid morphogenesis was studied by light microscopy through the analysis of meiotic and premeiotic lesions. Sperm disfunction-type male-sterile mutations were screened for novel “early effect” mutations: (1) timing mutations, in which mitochondrial aggregation occurs before instead of after meiosis; (2) mutations which affect the spindle structure, e.g., a mutant with second-division monoastral spindle; (3) mutations which cause deformations in primary spermatocyte structures. It is shown, in addition to the examples cited above, that normal meiosis may often serve as an early marker for normal differentiation, and that approximately 20% of male-sterile mutations are meiotic mutants. The role of the Y chromosome was reexamined. The interaction between Y factors and X -linked male steriles is in many cases additive, indicating that Y gene products are essential for normal development of the primary spermatocytes. Furthermore, XO males are shown to be extreme meiotic mutants. It is argued that spermatid morphogenesis is totally dependent on developmental processes in the primary spermatocyte stage. The relations among developmental processes in early spermatogenesis are discussed in terms of gene activity.

Sequence replication and banding organization in the polytene chromosomes of Drosophila melanogaster

The relative proportions of cloned DNA fragments from all known hierarchies of sequence organization in polytene and diploid chromosomes were compared. It was found that unique sequences of varying sizes and chromosomal locations are equally replicated in salivary gland chromosomes. Sequences of euchromatic polydisperse gene families are also replicated proportionately in polytene and diploid tissues. Perhaps the most significant finding is that the histone gene repeats, despite their normal banding organization, are under-replicated in the polytene chromosome of Drosophila melanogaster. However, the clustered and well-banded 5S genes are most likely equally replicated. It is therefore concluded that differential sequence replication plays no apparent role in either the assembly or morphology of a band; and likewise, the assembly of polytenic DNA into band units is not affected by either the local abundancy or arrangement of middle repetitive sequences. The likelihood that the clustered arrangement is an important factor in the selection of sequences for under-replication is discussed.

The developmental program of spermiogenesis in Drosophila: a genetic analysis

Publisher Summary This chapter discusses the genetic program for male gametogenesis in Drosophila and presents a rationale for the genetic analysis of this developmental process. Spermiogenesis has been considered an excellent subject for genetic analysis. It is easy to obtain a large selection of noncomplementing, viable, male-sterile mutations in every part of the genome. The genetic and molecular organization of the Y chromosome in the Drosophila species and the possible function of its fertility genes are examined. The Y chromosome accounts for about 12% of the Drosophila genome but contains less than 10 fertility genes. These genes or some of them, form species-specific chromatin structures, resembling amphibian oocyte lampbrush loops. Thus, a unique opportunity to relate chromosome function and structure is presented in the chapter. The difficulties are that the effect of Y chromosome mutations on spermiogenesis is mimicked by many non-Y genes or by chromosomal aberrations and that almost all of the chromosomes are composed of repeated sequences. The Y chromosome is not engaged in meiotic recombination and it is part of the under-replicated and undifferentiated chromocenter in the polytene chromosomes. In Drosophila , deletions for clusters of 5 S genes can be temperature-sensitive, and in fact spermiogenesis itself, even in the context of the fly system, is temperature-sensitive. Most Drosophila lines became male sterile above 28°C, when viability as well as female fertility is otherwise good. Most important, a deletion of the whole Y chromosome is temperature-sensitive.

Heterochromatin markers: arrangement of obligatory heterochromatin, histone genes and multisite gene families in the interphase nucleus of D. melanogaster.

Localization, as detected by in situ hybridization, of major heterochromatic blocks in interphase nuclei of larval brain and imaginal discs is reported. We conclude that the position of heterochromatic regions in interphase nuclei is correlated with their respective position in metaphase chromosomes and hence, independent of sequence recognition. Furthermore, chromocentral associations of X-, Y- or autosomal-based heterochromatin are not formed in these cells. Homologues do align in close proximity, but heterochromatin plays no role in this arrangement. Heterochromatin, and probably nucleoli, establish their membrane links in situ, and have no prefixed recognition sites. The most intimate association between homologous repetitive sequences was found in the histone locus, but no tendency for clustering was found among loci of multisite euchromatic gene families.

Genetic identification of dominant overproducing mutations: The beadex gene

SummaryA model system for the identification of presumptive overproducing mutations from among visible dominant mutations in D. melanogaster is described. An overproducing mutation is expected if a dominant mutation is readily reverted by gene deletion and if gene deletions suppress the expression of the original dominant mutantion in flies heterozygous for the deletion. The Beadex (1:59.4) mutations are shown to satisfy these requirements, since a Bx dominant mutations is reverted by induced deletion [Df(Bx)/+) is wild type], and is also suppressed in trans by such a deletion [(bx/Df)Bx) is wild type].In addition, all 13 mutations recovered as Bx reversions or suppressors were associated with recessive held up (hdp) mutations allelic inter se, but not allelic to any known hdp gene. One such hdp mutations does not function as an independent dominant suppressor of Bx, is not always associated with Bx deletion, and in the latter situation is readily separable from Bx. We suggest that it functions as a Bx deletion, and may therefore represent the structural gene which is cis-regulated by the overproducing Bx mutations.

The role of X-linked lethal and viable male-sterile mutations in male gametogenesis of Drosophila melanogaster: genetic analysis.

SummaryThe possibility that viable male-sterile mutations occur in vital genes and the role played by lethal mutations and viable male-steriles in male gametogenesis were studied. Five sterile loci were identified among the 30 most proximal vital loci of the X-chromosome and two of them were shown to be allelic with lethal mutations. Fertility tests on gynanders for nonautonomous lethal mutations proved that vital genes operate autonomously in male gonads, independently of their effect on somatic tissues. Fertility tests of ts lethals, shifted to the nonpermissive temperature after the TSP, showed that 40% of vital genes function in male gonads. It is further shown that about the same proportion of vital genes is operating in female gonads and that the two groups overlap by about 70%. The role of viable and lethal male sterile mutations in the control and regulation of male gametogenesis is discussed in detail.

Genes Controlling Chromosome Activity: the Role of Genes Blocking Y-Lampbrush Loop Propagation

Characterization of two more X-linked Y-affecting mutations in D. hydei is presented; ms(1)XL24 is less extreme than the previously described mutation, ms(1)XL2, but its general effect on the propagation of Y-lampbrush loops is similar qualitatively; ms(1)XL24 is located at the distal end of the X-chromosome and associated with a suppressor-of-white phenotype. The other X-linked sterile, ms(1)XL4, is the least extreme of the three mutations examined to date. It is recessive, located ca 2 c.o. units to the right of white, and causes different patterns of Y loop response. — From a comparison of the effects of the three single mutations, as well as from the analysis of the various doublesterile combinations, we surmise that: 1) if specific activator genes for the Y-lampbrush loops do exist, they are not likely to be located on the X-chromosome of Drosophila; 2) every lampbrush loop is composed of several functional repeats, each of which operates when unfolded; 3) the Y-affecting mutations arrest preferentially, but not exclusively or specifically Y-lampbrnsh loop propagation probably indirectly through their independent metabolic effects. Since the mutations are not specific for germ line cells, and exhibit various pleiotropic effects, especially with regard to viability, they should be considered tissue conditional lethals.

Heterochromatin Markers: A Search for Heterochromatin Specific Middle Repetitive Sequences in Drosophila

We sought for cloned sequences of middle repetitive (MR) complexity that mark obligatory heterochromatic regions. Total genome probes were employed in a differential screening procedure to recover X-specific, Y-specific and autosomal specific heterochromatic sequences. X- and Y-linked sequences were recovered in the same experiment. (Y-linked clones will be described elsewhere). All nine independent, non-identical X-specific clones were found to be partially homologous to one another and to type I rDNA insertion. No other X-specific Bam HI or HindIII clones were found. In situ hybridization to normal and inverted chromosomes revealed extensive homology in the heterochromatin spanning the nucleolus organizer (NOR) and the eu-heterochromatin junction. Eleven clones which are underrepresented in polytene chromosomes were selected in another differential screening. None was autosome-specific. Five were of nucleolar origin. Among them a presumptive type II 28SrDNA insertion sequence was clearly localized within the X-chromosome proximal heterochromatin in addition to the known localization of the X and Y nucleolar organizers. We mapped three clones to major sites on the Y chromosome and to secondary autosomal sites. The results are discussed with regard to the complexity of heterochromatin organization.

Genes controlling chromosome activity

Normal propagation of Y chromosome lampbrush loops was used as a screening tool in order to recover X-linked mutations controling Y chromosome activation. The nature of the most extreme mutationthus recovered, sterile (1) XL2, is described. It is a recessive gene mutation, readily mapped 2 cross over units distally to white. The mutation exerts its sterilizing effect by blocking normal unfolding of all Y lampbrush loops, but does not affect the unique shape of each diminutive loop. The degree to which a loop forming site is developed is partially temperature sensitive. It is independent however, on its map location or the dose of homologous as well as heterologous sites. It was provisionally concluded therefore that site response to the XL2 effect is a stage specific and not a quantitative one. The possible ways by which non homologous genes control Y chromosome activity are discussed.

The zeste-white interaction: induction and genetic analysis of a novel class of zeste alleles.

The recovery and analysis of a novel class of zeste mutations is described. zop6 and zop11 do not require two w+ genes for their expression unless the z+ gene is present. Analysis of genetic interactions among zeste alleles proved that zop mutations are strong antagonists of the z+ gene product. zop6 is readily reverted by X‐rays or by ethylmethane sulphonate to a range of intermediate zeste alleles and thus it is considered to be an insertional mutation. We suggest that z1 and op mutations arose as a result of insertions in a presumptive control region, and two alternative models for the structure of the zeste locus are evaluated.