male-specific lethal 2

DEVELOPMENTAL BIOLOGY

Embryonic

In embryonic nuclei, MSLs and monoacetylated histone H4 associate with the X chromosome. This was established using double fluorescent in situ hybridization with an X chromosome DNA probe and immunostaining for MSL protein. The late blastoderm/early gastrula are the earliest stages at which the MSLs are detected in the subnuclear localization pattern characteristic of male embryos. Earlier than this, MSLs are rather homogeneously distributed in the nucleus. It seems that MSLs become localized simultaneously, or over a very short period of time. MSL localization to the X chromosome without colocalization of monoacetylated H4 has been thought to suggest that MSLs localize prior to histone H4 (Franke, 1996).

Effects of Mutation or Deletion

There is no significant amount of embryonic death caused by msl-2 mutation, and no significant effect of msl-2 on viability of females or heterozygous males. Although most msl-2 males are killed in the larval stages, some do reach the prepupal stage. Mutants develop very slowly, most remaining in the third-instar stage for several days until dying as larvae or in the process of forming a puparium (Belote, 1980).

Msl-2 is required for the male-specific assembly of a dosage compensation regulatory complex on the X chromosome of Drosophila. Msl-2 binds in a reproducible, partial pattern to the male X chromosome in the absence of Mle or Msl-3, or when ectopically expressed at a low level in females. Moreover, the pattern of Msl-2 binding corresponds precisely in each case to that of MSL-1, suggesting that the two proteins function together to associate with the X chromosome. When MSL-1 function is compromised, Msl-2 is only localized to sites where the mutant form of MSL-1 is bound. Thus, a partial Msl-2 pattern is observed in msl-1 mutants. Likewise, when Msl-2 is present in limiting quantities, MSL-1 is precisely restricted to the sites of Msl-2 localization. Ectopic Msl-2 expression in females results in a dominant female phenotype. EMS-induced loss of function msl-1 and msl-2 alleles were isolated in a screen for suppressors of the toxic effects of Msl-2 expression in females. One such mutant lacks the RING finger motif. Site-directed mutagenesis was also used to determine the importance of the Msl-2 RING finger domain and second cysteine-rich motif. The mutations, including those in conserved zinc coordinating cysteines, confirm that the RING finger is essential for Msl-2 function, while suggesting a less stringent requirement for an intact second motif, the metalothionein-like cysteine cluster known as a PHD finger (Lyman, 1997).

The binding of each of the MSLs, and of histone H4 acetylated on lysine 16, to the X chromosome requires the presence of all four MSL proteins. For example, no association of the MSL-3 protein with the X chromosome is detected in msl-1 mutants. Throughout embryogenesis, in none of the homozygous male embryos mutant for msl-2 or msl-3 could association of the other MSL proteins or monoacetylated histone with the X chromosome be detected. msl-1 mutant embryos show association of MSL proteins with X chromosomes up to stage 9, while in mle mutants such association is seen up to stage 10, but this binding can be attributed to the presence of maternal MLE and MSL-1 (Franke, 1996).

In Drosophila, dosage compensation requires assembly of the Male Specific Lethal (MSL) protein complex for doubling transcription of most X-linked genes in males. The recognition of the X chromosome by the MSL complex has been suggested to include initial assembly at ~35 chromatin entry sites and subsequent spreading of mature complexes in cis to numerous additional sites along the chromosome. To understand this process further, MSL patterns were examined in a range of wild-type and mutant backgrounds producing different amounts of MSL components. The data support a model in which MSL complex binding to the X is directed by a hierarchy of target sites that display different affinities for the MSL proteins. Chromatin entry sites differ in their ability to provide local intensive binding of complexes to adjacent regions, and need high MSL complex titers to achieve this. A set of definite autosomal regions (~70), competent to associate with the functional MSL complex in wild-type males, was mapped. Overexpression of both MSL1 and MSL2 stabilizes this binding and results in inappropriate MSL binding to the chromocenter and the 4th chromosome. Thus, wild-type MSL complex titers are critical for correct targeting to the X chromosome (Demakova, 2003).

A functional dosage compensation complex required for male killing in Drosophila

Bacteria that selectively kill males ('male-killers') were first characterized more than 50 years ago in Drosophila and have proved to be common in insects. However, the mechanism by which sex specificity of virulence is achieved has remained unknown. This study tested the ability of Spiroplasma poulsonii to kill Drosophila melanogaster males carrying mutations in genes that encode the dosage compensation complex. The bacterium failed to kill males lacking any of the five protein components of the complex (Veneti, 2005).

Certain isofemale lines of Drosophila only give rise to daughters following the death of male embryos. Male death is due to the presence of intracellular bacteria that pass from a female to her progeny and that selectively kill males during embryogenesis. These male-killing bacteria are found in a wide range of other insect species, and many different bacteria have evolved male-killing phenotypes. In some host species, male-killers drive the host population sex ratio to levels as high as 100 females per male and alter the pattern of mate competition. However, the underlying processes that produce male-limited mortality are unclear. This study examined the interaction between the male-killing bacterium Spiroplasma poulsonii and the sex determination pathway of D. melanogaster (Veneti, 2005).

The primary signal of sex in Drosophila is the X-to-autosome ratio. This signal is permanently established in expression of Sex-lethal (Sxl) in females and its absence in males. This, in turn, effects three processes: germline sexual identity, somatic sexual differentiation, and dosage compensation, the process by which the gene expression titer on the X chromosome is equalized between two sexes despite their difference in X chromosome number. Mutations in the gene tra that convert XX individuals to male somatic sex do not induce female death. These observations indicate germline formation and migration happen correctly in male embryos and that dying male embryos do not express Sxl. Therefore, the requirement of the Spiroplasma for genes within the system of dosage compensation was examined (Veneti, 2005).

In Drosophila, the single X of males is hypertranscribed. This process of hypertranscription requires the formation of the dosage compensation complex (DCC) and its binding to (and modification of) the X chromosome. SXL in female Drosophila inhibits the production of MSL-2 protein, which is thus only present in male Drosophila. MSL-2 forms a complex with four other proteins, MSL-1, MSL-3, MLE, and MOF, which collectively form the DCC. MSL-1, MSL-3, MLE, and MOF are constitutively present in both males and females and are also supplied maternally. The complete DCC binds, with JIL-1, to the male X chromosome at various entry points, and, with the products of two noncoding RNAs, RoX1 and RoX2, it affects the modification of the single X chromosome and its hypertranscription (Veneti, 2005).

The effect was examined of mutations within the host DCC on the ability of the male-killer to function. The survival of male progeny beyond embryogenesis to L2/L3 (and in one case adult) was scored in the presence of different loss-of-function mutations within the dosage compensation system (normal male-killing occurs during embryogenesis), in the presence and absence of infection. Because many genes within this group additionally show strong maternal effects, the effect of mutations was in each case tested by using both mothers that were heterozygous for the loss-of-function mutation and mothers homozygous for it (Veneti, 2005).

Uninfected males homozygous for loss-of-function mutations within the dosage compensation system generally survive to the third larval instar. Survival to the third larval instar was studied for loss-of-function alleles of msl-1 (alleles msl-1¹ and msl-1^b), msl-2 (msl-2^g227 and msl-2^g134), msl-3 (msl-3¹³²), mle (mle⁹, mle¹), and mof (mof¹), and survival to adult for mle¹/mle⁶ transheterozygotes. In the case of all alleles of msl-1, msl-3, mle, and mof, a similar pattern is observed: Males homozygous or hemizygous for the loss-of-function mutation have appreciable survival in the presence of infection when their mother is also homozygous for the loss-of-function mutation. In contrast, heterozygous male embryos that were siblings of the above (that will have a wild-type DCC) were always killed, as were all male embryos in the case where the mother was heterozygous (where maternal supply of these proteins enables dosage compensation to be initiated, although not maintained). In the case of mle¹/mle⁶ transheterozygotes, male survival to adult was observed. For the case of mof, male-killing was restored to full efficiency when 18H1, a transgenic copy of mof, was added to the mof¹ loss-of-function background. Within the above crosses, three observations make sure the observation that Spiroplasma was fully operational. First, crosses involving heterozygous mothers, where male-killing was complete, were conducted concurrently with those using homozygous mothers, and the females in these crosses were siblings from the same vials. Second, in each cross and vial where homozygous males survived, heterozygous males (with wild-type function) still died. Finally, F₁ females derived from these crosses, when mated to wild-type males, produced a full, female-biased sex ratio (Veneti, 2005).

In the case of msl-2, where there is no maternal supply of MSL-2, survival of homozygous sons was observed for both homozygous and heterozygous mothers for the case of the mutation msl-2^g134. For the case of msl-2 ^g227, no male progeny were observed from infected females (table S5). This mutation does not rescue males in the assay, probably because the two mutations have different effects on msl-2 expression. The msl-2^g134 allele prevents formation of MSL-2 protein, whereas msl-2 ^g227 potentially encodes the RING-finger element of a truncated MSL-2 protein (Veneti, 2005).

Thus, absence or reduced function of any of the proteins of the DCC can reduce the efficiency of male killing, and a functional DCC is required for male killing by S. poulsonii. The fact that the genes mediating this process in Drosophila have been well studied can be exploited to yield further insights into the mechanism of male killing (Veneti, 2005).

Abaza, I., Coll, O., Patalano, S. and Gebauer, F. (2006). Drosophila UNR is required for translational repression of male-specific-lethal 2 mRNA during regulation of X-chromosome dosage compensation. Genes Dev. 20: 380-389. 16452509

Amrein, H. and Axel, R. (1997). Genes expressed in neurons of adult male Drosophila. Cell 88: 459-469

Bashaw,G.J.and Baker,B.S. (1995). The msl-2 dosage compensation gene of Drosophila encodes a putative DNA-binding protein whose expression is sex specifically regulated by Sex-lethal. Development 121: 3245--3258

Bashaw, G. J. and Baker, B. S. (1997). The regulation of Drosophila msl-2 gene reveals a function for Sex-lethal in translational control. Cell 89: 789-798

Beckmann, K., et al. (2005). A dual inhibitory mechanism restricts msl-2 mRNA translation for dosage compensation in Drosophila. Cell 122: 529-540. 16122421

Belote, J., and Lucchesi, J. C. (1980). Male-specific lethal mutations in Drosophila melanogaster. Genetics 96: 165-185

Bernstein, M. and Cline, T. W. (1994). Differential effects of Sex-lethal mutations on dosage compensation early in Drosophila development. Genetics 136: 1051-61

Bhadra, U., Pal-Bhadra, M. and Birchler, J. A. (1999). Role of the male specific lethal (msl) genes in modifying the effects of sex chromosomal dosage in Drosophila. Genetics 152(1): 249-268.

Bhadra, U., Pal-Bhadra, M. and Birchler, J. A. (2000). Histone acetylation and gene expression analysis of Sex lethal mutants in Drosophila. Genetics 155: 753-763.

Braunstein, M., et al. (1993). Transcriptional silencing in yeast is associated with reduced nucleosome acetylation. Genes Dev. 7: 592-694

Bone, J. R., et al. (1994). Acetylated histone H4 on the male X chromosome is associated with dosage compensation in Drosophila. Genes Dev. 8: 96-104

Buscaino, A., et al. (2003). MOF-regulated acetylation of MSL-3 in the Drosophila dosage compensation complex. Molec. Cell 11: 1265-1277. 12769850

Chang, K. A. and Kuroda, M. I. (1998). Modulation of MSL1 abundance in female Drosophila contributes to the sex specificity of dosage compensation. Genetics 150(2): 699-709.

Copps, K., et al. (1998). Complex formation by the Drosophila MSL proteins: role of the MSL2 RING finger in protein complex assembly. EMBO J. 17: 5409-5417.

Dahlsveen, I. K., Gilfillan, G. D., Shelest, V. I., Lamm, R. and Becker, P. B. (2006). Targeting determinants of dosage compensation in Drosophila. PLoS Genet. 2(2): e5. 16462942

Demakova, O. V., et al. (2003). The MSL complex levels are critical for its correct targeting to the chromosomes in Drosophila melanogaster. Chromosoma 112(3): 103-15. 14579126

Duncan, K., et al. (2006). Sex-lethal imparts a sex-specific function to UNR by recruiting it to the msl-2 mRNA 3' UTR: translational repression for dosage compensation. Genes Dev. 20: 368-379. 16452508

Forch, P., Merendino, L., Martinez, C. and Valcarcel, J. (2001). Modulation of msl-2 5' splice site recognition by Sex-lethal. RNA 7(9): 1185-91. 11565743

Franke, A., et al. (1996). Evidence that MSL-mediated dosage compensation in Drosophila begins at blastoderm. Development 122 (9): 2751-2760.

Franke, A. and Baker, B. S. (1999). The rox1 and rox2 RNAs are essential components of the compensasome, which mediates dosage compensation in Drosophila. Mol. Cell, 4: 117-122.

Friedman, J. R., et al. (1996). KAP-1, a novel corepressor for the highly conserved KRAB repression domain. Genes Dev. 10: 2067-78

Gebauer, F., et al. (1998). The Drosophila splicing regulator Sex-lethal directly inhibits translation of Male-specific-lethal 2 mRNA. RNA 4(2): 142-50.

Gebauer, F., et al. (1999). Translational control of dosage compensation in Drosophila by Sex-lethal: cooperative silencing via the 5' and 3' UTRs of msl-2 mRNA is independent of the poly(A) tail. EMBO J. 18: 6146-6154

Gebauer, F., Grskovic, M. and Hentze, M. W. (2003). Drosophila Sex-lethal inhibits the stable association of the 40S ribosomal subunit with msl-2 mRNA. Molec. Cell 11: 1397-1404. 12769862

Greenberg, A. J., Yanowitz, J. L. and Schedl, P. (2004). The Drosophila GAGA factor is required for dosage compensation in males and for the formation of the male-specific-lethal complex chromatin entry site at 12DE. Genetics 166: 279-289. 15020425

Grskovic, M., Hentze, M. W. and Gebauer, F. (2003). A co-repressor assembly nucleated by Sex-lethal in the 3'UTR mediates translational control of Drosophila msl-2 mRNA. EMBO J. 22: 5571-5581. 14532129

Gu, W., Szauter, P. and Lucchesi, J. C. (1998). Targeting of MOF, a putative histone acetyl transferase, to the X chromosome of Drosophila melanogaster. Dev. Genet. 22(1): 56-64. 9816055

Hamada, F. N., et al. (2005). Global regulation of X chromosomal genes by the MSL complex in Drosophila melanogaster. Genes Dev. 19: 2289-2294. 16204180

Hilfiker, A., et al. (1995). The gene virilizer is required for female-specific splicing controlled by Sxl, the master gene for sexual development in Drosophila. Development 121: 4017-4026. 8575302

Hilfiker, A., et al. (1997). mof, a putative acetyl transferase gene related to the Tip60 and MOZ human genes and to the SAS genes of yeast, is required for dosage compensation in Drosophila. EMBO J. 16(8): 2054-2060.

Jin, Y., et al. (1999). JIL-1: a novel chromosomal tandem kinase implicated in transcriptional regulation in Drosophila. Mol. Cell 4(1): 129-35. <>Jin, Y., et al. (2000). JIL-1, a chromosomal kinase implicated in regulation of chromatin structure, associates with the male specific lethal (MSL) dosage compensation complex. J. Cell Biol. 149(5): 1005-10

Kelly, R. L., et al. (1995). Expression of msl-2 causes assembly of dosage compensation regulators on the X chromosomes and female lethality in Drosophila. Cell 81: 867-877

Kelley, R. L., et al. (1997). Sex lethal controls dosage compensation in Drosophila by a non-splicing mechanism. Nature 387 (6629): 195-199.

Kelley, R. I. and Kuroda, M. I. (2003). The Drosophila roX1 RNA gene can overcome silent chromatin by recruiting the male-specific lethal dosage compensation complex. Genetics 164: 565-574. 12807777

Li, F., Parry, D. A. and Scott, M. J. (2005). The amino-terminal region of Drosophila MSL1 contains basic, glycine-rich, and leucine zipper-like motifs that promote X chromosome binding, self-association, and MSL2 binding, respectively. Mol. Cell. Biol. 25(20): 8913-24. 16199870

Lyman, L. M., et al. (1997). Drosophila male-specific lethal-2 protein: structure/function analysis and dependence on MSL-1 for chromosome association. Genetics 147(4): 1743-1753.

Marcand, S., et al. (1996). Silencing of genes at nontelomeric sites in yeast is controlled by sequestration of silencing factors at telomeres by Rap 1 protein. Genes Dev. 10:1297-1309.

Marin, I. (2003). Evolution of chromatin-remodeling complexes: comparative genomics reveals the ancient origin of 'Novel' compensasome genes. J. Mol. Evol 56: 527-539. 12698291

McDowell, K. A., Hilfiker, A. and Lucchesi, J. C. (1996). Dosage compensation in Drosophila: the X chromosome binding of MSL-1 and MSL-2 in female embryos is prevented by the early expression of the Sxl gene. Mech. Dev. 57 113-119

Meller, V. H., et al. (1997). roX1 RNA paints the X chromosome of male Drosophila and is regulated by the dosage compensation system. Cell 88: 445-457

Meller, V. H., et al. (2000). Ordered assembly of roX RNAs into MSL complexes on the dosage-compensated X chromosome in Drosophila. Curr. Biol. 10: 136-143

Meller, V. H. (2003). Initiation of dosage compensation in Drosophila embryos depends on expression of the roX RNAs. Mech. Dev. 120: 759-767. 12915227

Mendjan. S., et al. (2006). Nuclear pore components are involved in the transcriptional regulation of dosage compensation in Drosophila. Mol. Cell 21: 811-823. 16543150

Merendino, L., Guth, S., Bilbao, D., Martinez, C. and Valcarcel, J. (1999). Inhibition of msl-2 splicing by Sex-lethal reveals interaction between U2AF35 and the 3' splice site AG. Nature 402(6763): 838-41. 10617208

Oh, H., Park, Y. and Kuroda, M. I. (2003). Local spreading of MSL complexes from roX genes on the Drosophila X chromosome. Genes Dev. 17: 1334-1339. 12782651

Oh, H., Bone, J. R. and Kuroda, M. I. (2004). Multiple classes of MSL binding sites target dosage compensation to the X chromosome of Drosophila. Curr. Biol. 14: 481-487. 15043812

Park, Y., et al. (2003). Sequence-specific targeting of Drosophila roX genes by the MSL dosage compensation complex. Molec. Cell 11: 977-986. 12718883

Rattner, B. P. and Meller, V. H. (2004). Drosophila Male-specific lethal 2 protein controls sex-specific expression of the roX genes. Genetics 166: 1825-1832. 15126401

Scott, M. J., et al. (2000). MSL1 plays a central role in assembly of the MSL complex, essential for dosage compensation in Drosophila. EMBO J. 19: 144-155

Sobel, R. E., et al. (1995). Conservation of deposition-related acetylation sites in newly synthesized histones H3 and H4. Proc. Natl. Acad. Sci. 92: 1237-41

Straub, T., et al. (2005a). Stable chromosomal association of MSL2 defines a dosage-compensated nuclear compartment. Chromosoma 114(5): 352-64. 16179989

Straub. T., Gilfillan, G. C., Maier, V. K. and Becker, P. B. (2005b). The Drosophila MSL complex activates the transcription of target genes. Genes Dev. 19: 2284-2288. 16204179

Turner, B. M., Birley, S. J., and Lavender, J. (1992). Histone H4 isoforms acetylated at specific lysine residues define individual chromosomes and chromatin domains in Drosophila polytene nuclei. Cell 69: 375-384

Veneti, Z., Bentley, J. K., Koana, T., Braig, H. R. and Hurst, G. D. (2005). A functional dosage compensation complex required for male killing in Drosophila. Science 307(5714): 1461-3. 15746426

Zhou, S. Y., et al. (1995). Male-specific lethal 2, a dosage compensation gene of Drosophila, undergoes sex-specific regulation and encodes a protein with a RING finger and a metallothionein-like cysteine cluster. EMBO J. 14: 2884-95

date revised: 10 February 2007

 

Home page: The Interactive Fly © 1997 Thomas B. Brody, Ph.D.

The Interactive Fly resides on the
Society for Developmental Biology's Web server.