Science/Medicine : Sex and Fertility : By looking at the age-old mystery of gender selection, researchers hope to lay the groundwork to address currently untreatable chromosomal abnormalities that leave thousands sterile.
Open the “mammals” file on David Page’s computer and out spill charts and graphs bearing entries for a hodgepodge of creatures--goats, mice, chimps and humans--which have some chromosomal defect that has affected their sexual development.
And that information is helping Page answer a question that has long puzzled mankind: What determines sex?
Page, a geneticist with the Whitehead Institute for Biomedical Research here, is homing in on what geneticists call the “testes determining factor,” or TDF. His work is leading him slowly, inexorably along the familiar Y chromosome to a much smaller genetic unit, possibly even to a single gene.
“That,” said Page, “is the real source of excitement. Because here we have this age-old mystery that we have made many hypotheses about but have never had the tools to crack.”
By studying TDF, Page hopes to unearth clues that may lay the groundwork for addressing currently untreatable chromosomal abnormalities that render thousands of men and women sterile.
“Within a year or two, at the most, we may well have a reasonable idea as to the biochemical nature of this gene--whether it’s a single gene or whether it might be multiple genes,” he said. “Eventually we hope to understand the basis of infertility in these individuals.”
The question of sex determination, meanwhile, has inspired theories from the sublime to the silly.
“In the 1880s and ‘90s, there was a great deal of discussion into the impact of the parents’ diet on the sex of their child,” Page said. “And some of the ancient Greek philosophers thought that the sex of the child was determined by the level of excitement of the male partner during intercourse--the more excited the male partner, he greater the chances he would sire a son.”
Then early in this century, scientists learned that humans have 23 pairs of chromosomes, which contain all the body’s genetic material--with each parent contributing half of each pair at conception.
Among these pairs are the sex chromosomes, called X and Y. And it was determined that an embryo gets an X chromosome from its mother and an X or a Y from its father.
While it was known that an embryo with two X chromosomes became a female and an XY embryo became male, it wasn’t until the late 1950s that the presence or absence of the Y chromosome was identified as the determining factor in sex.
But the same sophisticated instruments that produced this discovery also revealed that there were exceptions: women with XY chromosomes and men with XX chromosomes. Scientists speculated that a sub-factor on the Y chromosome somehow directed a developing embryo to produce testicles. If this sub-factor were missing, XY females could result. Similarly, if it got onto an X chromosome, an XX male was possible.
Further research showed that approximately one in every 20,000 men is XX.
One young man who was following these developments as they unfolded was Page, then a medical student at Harvard University working under a special program in the Massachusetts Institute of Technology’s biology department.
“It was really a series of unexpected experimental observations, one after another, that led me to study this,” Page recalled. A key was the discovery of certain DNA sequences, called “probes,” that enabled him to compare different sex chromosomes.
“I then started thinking: Maybe we could use this probe of the X and Y chromosomes to begin studying chromosomal abnormalities,” he said.
A tall, friendly man who often roams his lab in Hawaiian shirts, Page by then was already earning a reputation as a brilliant researcher. And beginning in 1984, as a physician and a geneticist, he launched an all-out effort to pin down the TDF.
One problem he faced was that there is no easy way to detect the abnormalities.
While many XY women lack normal breast development or do not menstruate--indicating they have not gone through normal puberty--the XX men exhibit few outward signs. Such men tend to be taller and leaner than average, but otherwise seem ordinary; most discover their abnormality only after they see a doctor to learn why they cannot sire children.
So Page wrote to endocrinologists and clinical geneticists around the country asking for blood samples or skin biopsies of people with sex chromosome anomalies.
His work is a something of an oddity in biomedical research in that it is the 100 or so humans he studies that provide the basis for subsequent animal research. Typically, such research works the other way around.
“It’s precisely why these studies have gone so much further in humans than they have in mice,” Page explained. “People, unlike mice, complain about problems like infertility.”
In his human studies, Page begins with a set of cloned DNA sequences from a normal Y chromosome. The chromosomal DNA is made up of long double strands of nucleotides of four types--adenine, cytosine, guanine and thymine. The nucleotides bind together in what are called “complementary base pairs”: Adenine always binds with cytosine, and guanine always pairs with thymine.
It is the sequence of these nucleotides that makes up the genetic message, and these sequences are what Page examines in his research.
Using a technique involving restriction enzymes, which are known to cut through DNA at specific combinations of nucleotides, Page can remove fragments of the chromosome.
He then adds radioactively labeled DNA probes from normal chromosomes--some of the same ones he discovered while a medical student--that attempt to find their complementary nucleotides. If the two strands bind, Page knows that that part of the subject’s chromosome is present and normal.
By painstakingly charting his results, Page has narrowed the search to “something less than 1% of the Y chromosome.” He now suspects a single gene is responsible for TDF and is developing ever-more-specific probes to find it.
Last year he won a prestigious MacArthur Foundation fellowship, the so-called “genius award,” for his pioneering studies.
But Page concedes that much work lies ahead in tracking down the sex-determining gene. “It is precisely like looking for a needle in a haystack,” he said with a smile. “But the haystack is getting much smaller.”
Already, Page has learned that these chromosomal defects occur randomly, rather than as genetic mutations that are passed on. “Most of the things we study occur once in a family,” he said. “Very, very rarely do you see two XX males in a family.”
The abnormalities, he has determined, arise during sperm formation, when parts of the father’s own X and Y chromosomes pair and recombine in a process called “crossing over.” This exchange means that whatever sex chromosome the father passes to his offspring will contain genetic information from both of the father’s parents.
In the cases Page has studied, the wrong parts were swapped.
Thus, the sperm with the defective sex chromosome carries a piece of the father’s Y on the X chromosome, or vice versa.
During fetal development, the reproductive organs--the gonads--become either ovaries or testes, depending on the presence or absence of the Y chromosome, in particular the part that Page is searching for. This, in turn, lays the groundwork for sexual development by dictating the hormonal environment for all other processes in sex differentiation.
So why does sterility result?
“It appears you have to have an intact or nearly intact Y chromosome to be a fertile male,” Page explained. Similarly, he said, in females two intact or nearly intact X chromosomes are probably necessary.
Page is quick to point out that his work has no immediate practical implications for sterility. “But,” he said, “our studies have shed a lot of light on the genetic basis of sex determination.”
And once that mystery is solved, he and other scientists can begin to investigate the biochemical processes--involving hormones, structural proteins and enzymes--that ultimately determine both sex and fertility.
THE SEX DETERMINING FACTOR The Accepted Belief
Scientists have known for some time that humans have 23 pairs of chromosomes that contain all of the body’s genetic material, sequenced in strings of nucleotides known as DNA.
At conception, each parent contributes half of each pair of sex chromosomes--called X and Y. The X chromosome comes from the mother and an X or a Y from the father.
Generally speaking, an embryo with two X chromosomes becomes a female, and one with mixed chromosomes is male.
New research is now focusing on aberrant sexual development and sterility in an attempt to learn just how the genetic transfer occurs.
“Crossing Over”
During cell division, similar chromosomes--including sex chromosomes--normally pair up and exchange genetic material. The chromosomes overlap and adhere to each other--”crossing over” and exchanging sections.
The chromosome pairs split apart and each half contains the developmental information for separate cells.
The result of that cell division is a termed a gamete.
Page’s Belief
During sperm formation, part of the father’s own X and Y chromosomes pair and recombine in the “crossing-over” process. Geneticist David Page has focused his studies on the “testes determining factor” or TDF. He suggests that there is a genetic unit smaller than the Y chromosome--possibly even a single gene--that can code for maleness.
If that portion of a chromosome is inadvertently swapped during the crossing-over exchange, he contends, the normal development of reproductive organs may be altered.
There are, in fact, males whose cells show XX chromosome pairings and females who show XY pairings.
