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Figure 1 The paired X and Y chromosomes of themammalian spermatocyte form a uniquechromatin domain, marked here byimmunostaining for phosphorylated histone H2AX(green), which is not found in the autosomaldomain (red, immunostained for SYCP3, a proteinof the synaptonemal complex).

Imprinting is an epigenetic modification thatendows chromatin with a mark of parentalorigin1. In mammals, specific chromosomalregions, and whole chromosomes, aremarked as maternal or paternal. The nature ofthe mark is not fully understood, but bothmethylation of DNA and chromatin modifi-cations have been implicated. This epigeneticmark can result in differences in transcrip-tional activity, which is how many imprintingeffects are discovered. On page 100 of thisissue, Bean et al.2 provide a new twist to theimprinting story. They find that inCaenorhabditis elegans, an unpaired X chro-mosome acquires a chromatin imprint dur-ing gametogenic meiosis.

Singled outNematode sex is unorthodox. XX individualsare hermaphrodites and produce both spermand eggs, whereas XO individuals are com-mitted males, producing only sperm.Previous work3,4 showed dimorphism in thebehavior of the X chromosome in these twogerm lines. In both germ lines, the X chromo-some is inactive through most of meioticprophase. But in XX individuals, the X chro-mosome accumulates activating histonemodifications late in meiotic prophase duringoogenesis, whereas the univalent, unpaired Xchromosome in XO spermatocytes remainscondensed and inactive.

Bean et al.2 asked if these modificationsmight predict differences between the mater-nal and paternal X chromosome in XX zygotesproduced by mating XO males with XX indi-viduals. They assayed accumulation on chro-mosomes of histone modifications associatedwith transcriptional activation—dimethyla-tion of histone H3 at Lys4 and acetylation ofH3. In early embryos they found an excep-tional chromosome that did not have the his-tone modifications found on all otherchromosomes until after several rounds of celldivision. This chromosome was the paternallyderived X chromosome. No such exceptionalchromosome was found in the XO embryos,

whose sole X chromosome is maternallyderived. They found an X chromosome with-out activating histone modifications in thesperm pronucleus of self-progeny, but it accu-mulates activating histones earlier than doesthe X chromosome derived from XO males.

What could this mean? Obviously,germline sex is important since the paternalX chromosome differs from the maternal Xchromosome. But this is not the completestory, because the X chromosome from anXO male germ line has a more stableimprint than the paternal X chromosomefrom hermaphroditic (XX) male germ cells.So Bean et al.2 asked if the meiotic pairingstatus of the X chromosome might influ-ence the imprint, with an unpaired X chro-mosome acquiring a more stable imprint.They used the C. elegans sex determinationsystem and sex-reversing mutants to showthat pairing status of the X chromosome,more than germline sex, determines thechromatin imprint. The establishment ofthe imprint is probably related to accumu-lation on the unpaired X chromosome(both in XO males and in mutant XO her-maphrodites undergoing oogenesis) of his-tone H3 methylated at Lys9 (H3-Lys9), ahistone modification associated with thedevelopment of a repressive chromatinstructure5.

Silencing singlesMeiotic silencing is not a new observation. Ithas been well documented in mammalianspermatocytes6, where the X chromosomelacks a fully homologous pairing partner andtogether, the sex chromosomes assume aunique chromatin conformation (Fig. 1)known as the XY body that accumulatesmethylated H3-Lys9 (ref. 7) and many otherproteins. What probably set the authors downtheir current path of research is the knowledgethat transgene repeat arrays in C. elegans aresilenced in the germ line and accumulatemethylated H3-Lys9 (refs. 3,4). Meiotic silenc-ing by unpaired DNA (MSUD) is best studiedin Neurospora crassa8, where genetic analysishas implicated an RNA interference–mediatedsilencing9. MSUD might be a mechanism ofgenomic defense or protection against invad-ing sequences that has been hijacked by sexchromosomes in heterogametic species. But itcould pose an inconvenience for the species ifgenes required for spermatogenesis resided onthe X chromosome. In fact, the C. elegans Xchromosome is depleted of spermatogenesisgenes10 and silencing of the X chromosome inspermatogenesis may not be a problem.

But in mammals, there is a disproportion-ate abundance of spermatogenesis genes onthe X chromosome11. So what gives? Many ofthese genes are expressed only before11 orafter12 meiosis, circumventing the problem.But there are many housekeeping genes onthe mammalian X chromosome, and anothercoping strategy is implied by the existence ofautosomal paralogs that are expressed only inmeiotic prophase spermatocytes6. Perhapsthe X chromosome has not rid itself of thisinconvenient silencing because it is impor-tant for the paternal X chromosome to bemarked in the embryo. For instance, in XXmarsupials and mammals, there is nonran-dom inactivation of the paternal X chromo-some globally or in the trophectoderm.Although the paternal X chromosome waspreviously thought to be active in the zygote,new evidence suggests that XX zygotesinherit a preinactivated paternal X chromo-some13. For XY heterogametic sexes, howbetter to acquire a paternal X chromosomemark than by recognition of lack of pairingpartner?

12 VOLUME 36 | NUMBER 1 | JANUARY 2004 NATURE GENETICS

Marking Xs, together and separatelyMary Ann Handel

A chromosome imprinting mark is sensitive to meiotic pairing in the gamete of origin and identified through the‘histone code’.

Mary Ann Handel is at The Jackson Laboratory,600 Main Street, Bar Harbor, Maine, USA andthe University of Tennessee, Knoxville,Tennessee, USA. e-mail: mahandel@jax.org

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All this smacks of transvection and otherhomology-based silencing phenomena, whichare increasingly recognized as important devel-opmental mechanisms14. But the mark on thepaternal X chromosome might not be indelible,because male mice who inherit their X chromo-some from their fathers are not apparentlydevelopmentally impaired15. Notably, Bean etal.2 found that the imprint of the C. eleganspaternal X chromosome disappears after sev-eral rounds of mitosis as it accumulates histonemodifications. Although the observed histonemark disappeared, we don’t yet know if theunderlying imprint also disappeared.

Now the questions begin: how is lack ofpairing recognized, how is the imprint estab-lished and what is the underlying imprint?One thing is clear: the special status of the

unpaired X chromosome of XO males is iden-tified through the histone code. Althoughmany will assume that the special X chromo-some is transcriptionally silent, this has notyet been rigorously demonstrated. But what-ever the epigenetic mark and whatever itsfunction, knowledge of the roles of modifiedhistones will take us closer to discovery.

This paper provides grist for the experi-mental mill of many of us: those interested inmeiosis, reproduction, embryogenesis, tran-scriptional control, chromatin and evolution.It has wide implications to be tested in severalsystems, and I, for one, am quite eager to seewhere this takes us!

1. Reik, W. & Walter, J. Nat. Rev. Genet. 2, 21–32 (2001).2. Bean, C.J., Schaner, C.E. & Kelly, W.G. Nat. Genet.

36, 100–105 (2004).

3. Kelly, W.G. et al. Development 129, 479–492(2002).

4. Reuben, M. & Lin, R. Dev. Biol. 245, 71–82 (2002).5. Jenuwein, T. & Allis, C.D. Science 293, 1074–1080

(2001).6. McKee, B.D. & Handel, M.A. Chromosoma 102,

71–80 (1993).7. Peters, A.H.F.M. et al. Cell 107, 323–337 (2001).8. Shui, P.K. & Metzenberg, R.L. Genetics 161,

1483–1495 (2002).9. Lee, D.W., Pratt, R.J., McLaughlin, M. & Aramayo, R.

Genetics 164, 821–828 (2003).10. Reinke, V. et al. Mol. Cell 6, 605–616 (2000).11. Wang, P.J., McCarrey, J.R., Yang, F. & Page, D.C. Nat.

Genet. 27, 422–426 (2001).12. Moss, S.B., VanScoy, H. & Gerton, G.L. Mamm.

Genome. 8, 37–38 (1997).13. Huynh, K.D. & Lee, J.T. Nature advance online pub-

lication, 7 December 2003 (doi:10.1038/nature02222).

14. Wu, C. & Morris, J.R. Curr. Opin. Genet. Dev. 9,237–246 (1999).

15. Handel, M.A. & Hunt, P.A. Mol. Reprod. Dev. 28,337–340 (1991).

Diverse powerhousesDavid R Thorburn

Mitochondria in different tissues vary in number, morphology, ultrastructure, respiratory capacity and involvement inspecific metabolic pathways. A comparison of the proteome of mitochondria from different tissues has identified theextent of the underlying variation in protein composition and how this may be determined by tissue-specific networksof coregulated genes.

For an organelle that once seemed to be lovedonly by hard core biophysicists, the mitochon-drion has come a long way. Severe disorders ofmitochondrial oxidative phosphorylation(OXPHOS) are now recognized as the mostcommon group of inborn errors of metabolism,affecting at least 1 in 5,000 individuals1.Mitochondria are best known as the cell’senergy source as producers of ATP. But, theyalso have pivotal roles in generating reactiveoxygen species, calcium metabolism and celldeath. Thus, it is not surprising that mitochon-drial dysfunction contributes to diversepathologies including neurodegeneration, dia-betic complications and tumorigenesis. A recentstudy by Vamsi Mootha and colleagues2 in Cellprovides a basis for connecting mitochondrialpathologies with molecular etiology by identify-ing new mitochondrial proteins and profilingthe extent of their tissue-specific diversity.

David R Thorburn is at The Murdoch ChildrensResearch Institute and Genetic Health ServicesVictoria, Royal Children’s Hospital, andDepartment of Paediatrics, University ofMelbourne, Melbourne, Australia.e-mail: david.thorburn@mcri.edu.au

Compiling the listOver the past few years, different strategies

have been used in attempting to determinethe total number of proteins in mitochon-dria. These include epitope-tagging, system-atic functional screening of whole-genome

pools of mutants and proteomic analyses ofhighly purified mitochondria. Bioinformaticanalyses have also been used to predict pro-teins with a classical mitochondrial targetingsequence or genes that are coregulated withgenes encoding known mitochondrial pro-teins. These complementary approaches arenecessary to overcome the various limitationsin sensitivity and specificity of each method.Recent studies using such approaches in yeastare consistent with an estimate of 800–1,000different mitochondrial proteins3–6.

Yeast mitochondria are relatively simpleand uniform, but mammalian mitochondriavary widely between different tissues andoften don't resemble the text-book version(Fig. 1) in morphology or composition7

(Table 1). The mitochondrial proteome ofmammals is probably much larger andmore diverse than that of yeast, but its sizeand variability have not been ascertained.Computational analysis predicts up to 4,000mitochondrial proteins in humans3, whichmay be a true reflection of the complexity anddiversity of our mitochondria or an overesti-mate caused by a systematic artifact. The firstlarge-scale proteomic analysis of mammalianmitochondria, using human heart tissue, was

Figure 1 Colored high resolution scanningelectron micrograph of a single mitochondrion inthe cytoplasm of an intestinal epithelial cell.

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