Drosophila Cytogenetics Early Milestones

Sutton's 1903 landmark paper, "The Chromosomes in Heredity" (5; see also ref. 6), in which he pointed out that the behavior of chromosomes in meiosis parallels the observed patterns of inheritance of Mendelian traits, is considered to mark the beginning of the field of cytogenetics (the actual term would be coined years later). Before then, cytology with its focus on animals specimens, and genetics, which consisted of breeding experiments involving mainly plants, had been separate areas of inquiry (6). In the practical melding of cytology and genetics that soon followed, D. melanogaster would be unrivaled in its fundamental contributions to the new field of cytogenetics, many of which are listed in Table 1.

Ahead of her time and seldom acknowledged since, Nettie Stevens of Bryn Mawr College, Pennsylvania (34,35), was the first person to study chromosomes of Drosophila, beginning in the autumn of 1906 (7). Stevens, a codis-coverer of chromosomal sex determination, was already an accomplished cytologist when she worked the preceding summer at Cold Spring Harbor, NY, examining chromosomes of cucumber beetles (Diabrotica spp.; see ref. 36). It is then and there that she likely obtained her first Drosophila specimens from entomologist Frank Lutz. It was Lutz also, according to Kohler (37), who probably introduced Morgan to Drosophila that same year. Attempts in Morgan's laboratory to identify Drosophila mutations did not begin in earnest until either the fall of 1907 or 1908, and it was not until 1910 that the first unequivocal mutants were actually found (8,37). Thus, Stevens' cytological studies of D. melanogaster (then called D. ampelophila) predate the first use of Droso-phila for genetic analysis.

Stevens' research was important because it helped substantiate Sutton's chromosome theory of heredity (5). Indeed, she was far ahead of Morgan in recognizing the significance of chromosomes (34,35). For example, Stevens discovered that the karyotypes of male and female Drosophila (and other insects) differ at a single chromosome pair, from which she inferred a role for such heteromorphic chromosomes in determining sex (7,38), following

Table 1

Some Notable Achievements in Studies of Drosophila Chromosomes

1906 N. M. Stevens begins first ever study of Drosophila chromosomes;

observes heterochromosomes (X and Y) in males; observes somatic pairing of homologous chromosomes (7; see text).

1910 T. H. Morgan discovers sex-linked inheritance; first assignment of a specific gene (white) to a specific chromosome (the X) (8).

1913 A. H. Sturtevant constructs the first genetic map (involving X-linked genes) (9).

1914, 1916 C. W. Metz builds on Stevens' work; examines chromosomes in approx 80 species of Diptera, in gonads of both sexes, and in somatic tissues of embryos, larvae and pupae (10,11).

1916 C. B. Bridges proves chromosome theory of heredity through observa tions of nondisjunction (12).

1916 H. J. Muller discovers crossover suppressors, later shown to be chromo some inversions, from which the concept of "balancer" chromosome is derived (13).

1917, 1919 C. B. Bridges describes the first chromosome deficiency, first chromosome duplication (inferred from genetic analysis) (14,15).

1930 H. J. Muller discovers variegating mutations ("eversporting displacements," e.g., white-mottleds) resulting from chromosome rearrangements (16).

1930s E. Heitz investigates Drosophila heterochromatin (17-19).

1931 T. S. Painter discovers giant chromosomes in larval salivary glands and demonstrates their usefulness for mapping (20-25).

1934 B. P. Kaufmann publishes survey of chromosomes from various tissues of Drosophila; reports that cells of the larval brain are most useful for observing mitoses; presents extensive morphological description of same (26).

1935 C. B. Bridges devises map coordinate system for polytene chromosomes (27).

1938 H. J. Muller and colleagues coin the term "telomere" to describe the specialized ends of chromosomes (28).

1940s T. O. Caspersson undertakes first cytochemical studies using micros copy (29).

1959 K. W. Cooper undertakes extensive cytological investigation of hetero-

chromatin, including morphological descriptions of the X and Y hetero-chromatic elements (30).

1969 M.-L. Pardue and J. G. Gall develop in situ hybridization method for polytene chromosomes (31).

1972 D. L. Lindsley et al. systematically analyze the genome using synthetic duplications and deficiencies created in crosses of Y-autosome translocation stocks (32).

1977 G. T. Rudkin and B. D. Stollar demonstrate first FISH experiment (33).

Note: Some of these advances may be considered purely "genetic" rather than "cytogenetic," but they are included as historical reference points.

McClung (39). [N.B.: It was later established by Bridges that sex in Droso-phila is determined by the ratio of X chromosomes to sets of autosomes (40), and not by the presence or absence of a Y chromosome, as in humans, for example. The Y chromosome of Drosophila is essential for male fertility but not for maleness.] Stevens was also the first to note the tendency of homologous chromosomes of Diptera to pair in diploid somatic cells (i.e., outside of meiosis; see ref. 7).

The methods Stevens used to study insect chromosomes are not so different from the basic techniques we use today. She dissected testes and ovaries of adult flies in physiological salt solution, transferred the tissues to a drop of stain (acetocarmine) on a microscope slide, pressed the cover slip down to break and spread the cells, and removed the excess stain by wicking with filter paper. Of the nine dipteran species she studied, Stevens found the tissues of Drosophila to be the most difficult to work with, requiring her to examine an inordinate number of specimens. She wrote,

While in Sarcophaga all the stages necessary for a description of the behavior of the heterochromosomes of both sexes were found in the course of a few hours' work on perhaps ten or twelve preparations, satisfactory results in the case of Drosophila have been obtained only after prolonged study extending over more than a year and involving dissection of some two thousand individuals. (7)

(See Chapters 2-5, and reduce the number of your Drosophila dissections to Sarcophagan levels!) Despite such inauspicious beginnings, Stevens' camera lucida drawings of Drosophila prophase figures clearly show a complement of eight chromosomes, with males having a heteromorphic pair (later designated X and Y). Stevens concluded, "The general results of the nine species of flies are the same; i.e., an unequal pair of heterochromosomes in the male leading to dimorphism of the spermatozoa, and a corresponding equal pair in the female, each equivalent to the larger heterochromosome of the male. . ." (7). Unfortunately, Stevens' promising work on flies was abruptly stopped by the breast cancer that claimed her life in 1912. It was left to Charles Metz (10,11) and many others to build on Stevens' discoveries (see Table 1).

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