This is old Lecture 17.
Revised September 17, 1998
Textbook assignment: Chapter 9, pages 221-234 (to start of "Aneuploidy")
Major concepts
Chromosomal Sex Determination: This lecture deals primarily with the role of chromosomes in sex determination, with emphasis on humans and other mammals, where the presence of a Y chromosome is the primary determinant of maleness, and on Drosophila, where sex is determined by the ratio of X chromosomes to autosomes. The phenomenon of dosage compensation, which functionally equalizes the ratio of sex-linked genes to autosomal genes in males and females is also explored, together with an analysis of the effects of altered numbers of X and Y chromosomes on human sexual differentiaiton. I have chosen to introduce these topics well ahead of their position in the textbook in order to be able to discuss sex linkage (our next lecture) in terms of the numbers and behaviors of sex chromosomes found in males and females of various species. In order to understand this material, we will also have to cover a bit of the general material on human chromosome variations at the beginning of chapter 9.
Human chromosome number: The textbook provides a brief history of attempts to determine accurately the number of human chromosomes. It was not until 1956 that the correct number of 46 was finally established. For many years before that, the number was generally believed to be 48. The difficulty in determining the correct number arose from the fact that all of the chromosomes are packed tightly together in mitotic and meiotic cells such that in ordinary histological preparations, they cannot be clearly seen as individuals and counted.
Hypotonic swelling: The key to finally identifying the correct number of human chromosomes was a technique for mitotic arrest of the cells with colchicine (which disrupted the spindle fibers), followed by carefully controlled hypotonic swelling that was sufficient to separate the chromosomes without bursting the cells. The cells were then ruptured very gently, so that the chromosomes from each individual cell would settle into a well separated pattern on a microscope slide without losing their identity as a clearly delineated set. This allowed the chromosomes to be counted precisely and arranged in pairs as show in in Figure 2.5 in our textbook.
Karyotype analysis: Figure 2.5 quite accurately depicts the way in which karyotype analysis (the process of sorting out of individual chromosomes and deciding whether any are missing, duplicated, or morphologically altered) was originally done, based entirely on relative sizes of the chromosomes and positions of their centromeres. The subsequent development of staining techniques that produce banded patterns, such as those in Figure 9.1, has made the identification of individual chromosomes far easier and more precise. Notice, however, that the basic scheme for arrangement of the 22 pairs of autosomes plus the sex chromosomes has not changed.
Aneuploidy: The term euploid refers to having an exact multiple of a haploid set of unaltered normal chromosomes (including expected alternative sex chromosome patterns in species where sex is chromosomally determined). Any pattern that does not satisfy that criterion is referred to as aneuploid. Loss of one chromosome, resulting in one of the chromosomes being unpaired, is referred to as monosomy. The presence of an extra chromosome, causing one of the normal chromosomes to be present in three copies, is called trisomy. Changes in the structure of individual chromosomes that do not alter the total number of chromosomes also cause a karyotype to be considered aneuploid. Table 1 in the text introduces a variety of additional terms used to describe euploid and aneuploid variations in chromosome number.
Nondisjunction: The most common cause of monosomy and trisomy is nondisjunction, which was briefly introduced in Chapter 2 (page 39, Figure 2.14). During meiosis, homologous chromosomes pair, followed by random assortment of one member from each pair to each of the poles during the first division and separation of sister chromatids during the second division. This process normally generates gametes that contain precisely one haploid set of chromosomes. If disjunction fails to occur and both homologs (or sister chromatids) of one chromosome move to the same pole, the resulting gametes will lack one chromosome or have a duplicate copy of one of the chromosomes. When the abnormal gametes produced by nondisjunction fuse with normal gametes during fertilization, the resulting progeny will have 2n - 1 chromosomes (monosomy) or 2n + 1 chromosomes (trisomy).
Sex chromosomes: The concept of chromosomal differences between males and females was introduced briefly in Chapter 2 (pages 23-25). In humans and most mammals and also in Drosophila, females have two homologous X chromosomes, whereas males have only one X chromosome, paired with a much smaller Y chromosome that carries relatively little genetic information. In species with XX females and XY males, the male is referred to as the heterogametic sex because he produces two different kinds of haploid gametes, one containing an X chromosome and the other containing a Y chromosome. The female, whose haploid gametes all contain the same kind of sex chromosome, is referred to as the homogametic sex. Since all female gametes contain X chromosomes, the sex of the progeny is determined by whether fertilization is with a sperm cell containing an X chromosome to produce a female or a Y chromosome to produce a male. Despite the similarity of the XX/XY patterns, the actual molecular genetic mechanisms that are involved in determining female or male development occurs are very different in humans and other mammals than in Drosophila. We will begin with mammals, but before doing so it will be helpful to consider dosage compensation (which is covered a bit later in the textbook)
Dosage compensation in mammals: Gene dosage must be balanced quite precisely in higher organisms. An elaborate mechanism has evolved to insure that the ratio of functional X-chromosomal genes to autosomal genes remains constant despite a two-fold difference in X:A ratios between male and female mammals (including humans). Early in the development, there is random inactivation of one of the X chromosomes in each cell of female embryos. Throughout the rest of that female's lifespan in all of her cells other than her germ cells, the inactivated X chromosome remains partially condensed as a heterochromatic Barr body in the interphase nucleus (Figure 9.2). Its genes are not transcribed and its replication is delayed, occurring at the very end of the S period of the cell cycle. When abnormal numbers of X chromosomes are present, as in the aneuploidies described below, the number of Barr bodies is always one less than the total number of X chromosomes (Figure 9.3)
Functional hemizygosity in individual cells in females: The term hemizygous refers to having just one copy of a gene or group of genes in an otherwise diploid background. The classic example is a sex-linked gene carried on the X-chromosome, which is present in only one copy in XY males. This allows a recessive gene that is present only in a single copy to express its phenotype in the same manner as it does when it is in the homozygous state in an XX female. As a result of random inactivaiton of one of the X chromosomes in every cell, a female mammal is a mosaic of two cell types, one expressing the maternal X chromosome and the other expressing the paternal X chromosome (Fig. 9.4). This is evident, for example, in the orange and black pigmented areas of a calico or tortiseshell cat (Figure 9.5), where alternative pigmentation alleles are expressed in patches that represent the mitotic descendants of single cells with one or the other X chromosome active. A similar pattern also occurs in human females who are heterozygous for sex-linked anhydrotic ectodermal dysplasia (absence of sweat glands)(Fig. 9.6). Although no pigmentation is involved, patches of skin lacking and containing sweat glands are randomly distributed over the body surface.
Mechanism of X-chromosome inactivation: The mechanism responsible for X-chromosome inactivation remains incompletely understood. The process is controlled from a single region on the X-chromosome, called the X-inactivation center (XIC). That region contains a gene that codes for a rather large RNA that is not translated into a protein and is only transcribed from inactivated chromosomes. This gene is called XIST (for X-inactive transcript) in humans and Xist in mice. The XIST RNA appears to be directly involved in the inactivation process on each chromosome that is inactivated. There is no genetic expression in the XIC region of the chromosome that remains active, but the details of the switching mechanism remain somewhat obscure.
Human sex chromosome aneuploidies: Despite their major structural differences, X and Y chromosomes contain pseudoatosomal regions that permit them to come together as "homologous pairs" during meiotic prophase and to be independently assorted to the poles during the first meiotic division in much the same manner as autosomes. Like other kinds of monosomies and trisomies, sex chromosome aneuploidies can originate either during the first or second meiotic division (Fig. 2.14). Because dosage compensation inactivates all but one X chromosome in each cell, aneuploidies that increase or decrease the expected number of X chromosomes have relatively mild effects in humans and other mammals. However, because not all of the genes on the X-chromosome are inactivated, and because germ cells that do not contain the correct number of X-chromosomes tend to degenerate, there are definite syndromes associated with each class of X-chromosome aneuploidy.
Turner syndrome: Humans with one X and no Y chromosomes (XO) develop as females who exhibit characteristic features known as Turner syndrome. Their ovaries degenerate and fail to produce the hormones needed for sexual maturation. In addition, they are short in stature and often have thickened and webbed necks and a variety of other characteristic phenotypic traits (Fig. 9.1b). However, their intelligence is not affected and they can lead realtively normal lives, particularly when given hormonal therapy to achieve additional growth and sexual maturation. XO females exhibit no Barr bodies because they do not have a second X chromosome to be inactivated.
Kleinfelter syndrome: Humans with two X and one Y chromosome develop as males, but are sterile and often exhibit some female characteristics, such as enlarged breasts (Figure 9.1a). They are also generally mildly mentally retarded. The XXY state is known as Kleinfelter syndrome. It is most easily diagnosed by the presence of a Barr body in a male.
Metafemales: Humans with 3X chromosomes (XXX) develop as females who are phenotypically variable, ranging from fully fertile and normal appearing to sterile with amenorrhea and mild mental retardation. The XXX state is sometimes referred to as metafemale and is characterized by the presence of two Barr bodies.
XYY males: Humans with one X and two Y chromosomes (XYY) develop as normal fertile males, although they are usually taller than average. At one time, it was thought that they had strong criminal tendancies, but further studies have cast serious doubt on the original conclusions. There appears to be a slight excess of XYY males in penal instutions compared to the general population, but the vast majority of XYY men lead normal lives.
Genes on the Y chromosome determine maleness in mammals: In humans, individuals with one X chromosome and no Y chromosomes develop as females (although with some defects). Individuals with two X chromosomes and one Y chromosome develop as males (again with some defects). Thus, the default pathway of development in the absence of active signals appears to be female, with maleness being determined by the presence of a Y chromosome.
Mammalian sex-determining genes: The Y chromosome has been shown to carry a gene designated Sry in mice and SRY in humans, whose gene product is a transcription factor that causes the primitive gonad to develop into a testis. Two testicular hormones are responsible for diverting development of the rest of the reproductive system from the female pattern to the male. Mullerian inhibiting factor (MIF) prevents development of early primordia known as Mullerian ducts into the female oviducts and uterus. Testosterone promotes development of all of the internal and external male reproductive structures other than the testes.
Testicular feminization: One of the most phenotypically spectacular human mutations (which our textbook does not discuss) causes loss of function of testosterone receptors, which results in a syndrome known as testicular feminization. XY males that are carry this mutation on their X-chromosomes are totally unable to respond to testosterone and develop as phenotypic females. They lack oviducts and uterus because development of these structures is inhibited by MIF, and they also have undescended abdominal testes. However, their appearance is fully female, including normal female breast development as they mature. One of few external clues is their extreme lack of body hair, indlucing pubic hair (the hair on their heads is not affected). In some cases, their overall external phenotypes are so completely female that the condition is not fully diagnosed until the individuals go to fertility clinics as married women seeking to find out why they are infertile and have never menstruated.
X:A ratio determines sex in Drosophila: In Drosophila the Y chromosome has no effect on the sex of individuals, which is determined by the ratio of X chromosomes to haploid sets of autosomes (all of the chromosomes except sex chromosomes). A ratio of approximately 1.0 (studied in individuals with extra sets of autosomes or sex chromosomes) results in female development, whereas a ratio of approximately 0.5 results in maleness, with intersex development at intermediate ratios (Fig. 9.7).
Sex-determining genes in Drosophila: Recent analysis of molecular mechanisms has shown that the product of a gene designated Sxl acts as a master switch for a gene regulatory cascade that determines whether the male or female developmental pathways are activated. Sxl stands for "sex lethal" and was originally named that because loss of function mutations at this locus result in total absence of homozygous female progeny, with no effect on males. It is now known that production of the Sxl gene product is required for female development, and that a regulatory element in its promoter is sensitive to the ratio of regulatory proteins produced from genes on the X chromosome (numerator elements) and on autosomes (denomenator elements) respectively. The Sxl gene product in turn regulates a downstream cascade of gene expression changes that will be analyzed in greater detail in the developmental biology course, MCDB 4650. (Our textbook provides a partial summary of the cascade on pages 231-232, which you do not have to learn at this time.)
Dosage compensation in Drosophila: Dosage compensation between autosomal and X-linked genes also occurs in Drosophila, but with a totally different mechanism than in mammals. Transcription from the single X chromosome in male Drosophila occurs at roughly twice the rate of transcription from each of the two X-chromosomes in female Drosophila. (The textbook briefly summarizes the molecular mechanisms that are thought to be involved, but they also are not a required part of this course). Thus, in Drosophila, dosage compensation is achieved by adjusting the rate of transcription of genes on the X-chromosome, whereas in mammals, it is achieved by almost totally inactivating one of the X-chromosomes.
Gynandromorphs: Although relatively rare, loss of individual chromosomes can occur in mitotic cells. When one X chromosome is lost from one of the nuclei during early development of a female Drosophila, a portion of the body can have an X:A ratio of 0.5, while the remainder has an X:A ratio of 1.0. This can result in the development of a fly in which one side of the body is basically male while the other side is basically female. In addition, XX embryos that are heterozygous for sex linked genes, will sometimes display the dominant phenotype in the female half of the mature fly and the recessive phenotype in the male half (Fig 9.8).
Alternative sex chromosome systems: In other species, there are diverse patterns of chromosomal and non-chromosomal sex determination. In birds and some reptiles, as well as moths, the male is homogametic (ZZ) and the female heterogametic (ZW). In certain types of insects, females are XX and males are XO (with O indicating absence of a second sex chromosome). In bees, females are diploid and males are haploid. The small nematode, C. elegans, which has become a popular subject for research on developmental genetics, exhibits a similar pattern, except that worms with two X chromosomes are true hermaphrodites (with male and female in the same individual), while those with only one X chromosome are true males. In some species, sex is determined by environmental factors. In many plant species, as well as some animal species, both sexes may be present in the same individual. Our textbook does not describe these alternative systems of sex determination and we will not have much opportunity to explore them, but you need to be aware that there are far more patterns of sex determination than we have covered in this course.