|MadSci Network: Genetics|
Dear Sam, Due to a phenomenon known as imprinting, explained in more detail below, mammalian cloning does not work in the sense that most cloned animals are abnormal. Some people would say that all cloned mammals are abnormal. So the first task in a science fiction story about cloning is to figure out how to defeat imprinting. As gametes (sperm and ova) are formed, males and females "mark" known segments of the genome with an "imprint" that allows cells to distinguish whether they were inherited maternally or paternally. During the course of embryonic development, some tissues preferentially express the maternally-inherited gene, while other tissues do the reverse. In cloning, a nucleus from a somatic cell is transplanted to an enucleated egg. The challenge facing this nucleus is that since it has not been reprogrammed through gametogenesis in the male and female parents, it is not fully capable of directing normal development in the embryo derived from nuclear transfer. Here are some reviews on imprinting, often discussed in the context of cloning: Nat Rev Genet. 2001 Jan;2(1):21-32. N Engl J Med. 2003 Jul 17;349(3):211-2. Science. 2001 Aug 10;293(5532):1093-8. The last article is available online at: http://www.sciencemag.org/cgi/content/full/293/5532/1093 Compared to this barrier to cloning, sex reversal is rather straightforward. There are two possibilities. First, it would be possible (although unethical) using easily imagined genetic intervention to transform a male embryo into a phenotypic female incapable of reproduction. Second, it might be possible to perform a kind of genetic surgery on a male donor's nuclei to create a nucleus in which the Y chromosome was removed, and a second copy of his X chromosome was inserted. The second case is more easily explained. Imagine that we are able to identify and remove an X chromosome from a cultured cell from a male donor. Methods for "sorting" chromosomes using flow cytometry have been described. See, for example: Methods Enzymol. 1987;151:252-67. Methods Cell Biol. 1994;42 Pt B:319-30. Methods Mol Biol. 1994;29:205-19. It is not hard to imagine that we might be able to remove a Y chromosome from a cell and substitute a second copy of the X chromosome. We would still face the problem of getting this nucleus to serve as the basis of a successful clone by nuclear transfer as discussed above. If this procedure were successful, we could produce a female from a male donor. This female clone would be genotypically identical to the male donor except for the absence of the Y chromosome. In the other case that I raised, we can imagine directing development of a male embryo to be female, because the pathway for sex determination and differentiation in mammals is well known. A particular gene on the Y chromosome called SRY is key to male development. Early embryos are not sexually differentiated, having a presumptive gonad that is not clearly ova or testis, and the rudiments for all of the associated structures in both males and females are present as a set of ducts, the Mullerian and Wolffian ducts. If SRY is expressed in the presumptive gonad, differentiation is directed toward maleness, testosterone is produced, and the Wolffian duct differentiates into the epididymus, vas deferens, and seminal vesicle, while the Mullerian duct degenerates. If SRY is not expressed, development is female, the Mullerian duct becomes the uterus, oviduct, cervix and upper vagina, while the Wolffian duct degenerates. It would be possible to interfere with SRY expression using antisense RNA or targeted mutation, producing a person who is chromosomally male but who is phenotypically female. Such a person would be like chromosomal males with mutations in SRY. These individuals are described in OMIM at: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=OMIM Search on SRY to get a description of the gene or on Swyer syndrome. These individuals are not normal females; they do not develop secondary sexual characteristics at puberty. Another kind of inherited sex reversal results from inherited mutation in the androgen receptor. Search OMIM for Androgen Insensitivity Syndrome. Because the signals directing embryonic development to maleness are in part hormonal, inability to respond to androgen because of mutation in the receptor will cause XY males to develop as females. These individuals develop some secondary sexual characteristics at puberty but do not menstruate. They are infertile. It would be possible to create targeted mutations in the androgen receptor in a male donor nucleus to create a clone that was apparently female, but not fully so. This answer would not be complete without a brief discussion of ethics. There is no clinical reason to clone a person, especially given today's technology which virtually assures that cloned individuals will be abnormal. It is not possible to justify a medical procedure that results in the birth of an abnormal individual, especially because there is no benefit to be gained from this technique to the clone, the donor, or anyone else. If we were able to clone people with the assurance that they would develop normally, it is not possible to justify a medical intervention that reverses the sex of the clone either fully (using chromosome replacement) or partially (using mutations in the sex determination pathway). Having said that, there are some interesting ideas that could be explored in the literary, rather than medical, realm. Good luck with your story. Please see the MGI Glossary: http://www.informatics.jax.org/mgihome//other/glossary.shtml Yours, Paul Szauter Mouse Genome Informatics
Try the links in the MadSci Library for more information on Genetics.