InteractiveFly: GeneBrief

Heterochromatin Protein 1e: Biological Overview | References

Sperm-packaged DNA must undergo extensive reorganization to ensure its timely participation in embryonic mitosis. Whereas maternal control over this remodeling is well described, paternal contributions are virtually unknown. This study shows that Drosophila melanogaster males lacking Heterochromatin Protein 1E (HP1E) sire inviable embryos that undergo catastrophic mitosis. In these embryos, the paternal genome fails to condense and resolve into sister chromatids in synchrony with the maternal genome. This delay leads to a failure of paternal chromosomes, particularly the heterochromatin-rich sex chromosomes, to separate on the first mitotic spindle. Remarkably, HP1E is not inherited on mature sperm chromatin. Instead, HP1E primes paternal chromosomes during spermatogenesis to ensure faithful segregation post-fertilization. This transgenerational effect suggests that maternal control is necessary but not sufficient for transforming sperm DNA into a mitotically competent pronucleus. Instead, paternal action during spermiogenesis exerts post-fertilization control to ensure faithful chromosome segregation in the embryo (Levine, 2015).

Faithful chromosome segregation requires careful orchestration of chromosomal condensation, alignment, and movement of mitotic chromosomes during every eukaryotic cell division. The very first embryonic mitosis in animals requires additional synchronization. Paternally and maternally inherited genomes undergo independent chromatin reorganization and replication prior to mitotic entry. For instance, maternal chromosomes must complete meiosis and then transition from a meiotic conformation to an interphase-like state in preparation for replication. The sperm-deposited, paternal chromosomes must undergo an even more radical transition from a highly compact, protamine-rich state to a decondensed, histone-rich state before DNA replication. Despite these divergent requirements to achieve replication- and mitotic-competency, maternal and paternal genomes synchronously enter the first mitosis. Failure to carry out paternal chromosome remodeling in a timely fashion results in paternal genome loss and embryonic inviability (Levine, 2015).

The transition from a protamine-rich sperm nucleus to a competent paternal pronucleus requires the action of numerous maternally deposited proteins in the egg (McLay and Clarke, 2003). For instance, paternal genome decondensation post-fertilization requires the integration of histone H3.3, a histone variant deposited by the maternal proteins HIRA, CHD1, and Yemanuclein (Orsi, 2013). Similarly, maternally-deposited MH/Spartan protein localizes exclusively to the replicating paternal genome and is required for faithful paternal chromosome segregation during the first embryonic division (Delabaere, 2014). These and other studies demonstrate the essential role of maternally-deposited machinery in rendering competent sperm-deposited DNA and ultimately, ensuring faithful paternal genome inheritance (Levine, 2015).

Is paternal control also necessary for the extensive decondensation and re-condensation of the post-fertilization paternal genome? If so, disruption of such control would manifest as paternal effect lethality (PEL). Unlike male sterility mutants that lack motile sperm, PEL mutants make abundant motile sperm that fertilize eggs efficiently. However, embryos 'fathered' by PEL mutants are inviable. Only a handful of PEL genes have been characterized in animals. These encode proteins that mediate sperm release of paternal DNA, sperm centriole inheritance, and paternal chromosome segregation. Only one of these PEL proteins directly localizes to paternal chromosomes; the sperm-inherited K81 protein localizes exclusively to paternal chromosome termini and ensures telomere integrity. The maintenance of telomeric epigenetic identity joins a growing list of examples of sperm-to-embryo information transmission via protein or RNA inheritance (e.g., diet, stress, embryonic patterning, transcriptional competency). Despite a new appreciation of paternal control over epigenetic information transfer, there are no reports of paternal control over the global chromatin reorganization required for synchronous mitosis across paternally and maternally inherited genomes. Indeed, in the absence of any known paternal protein-directed genome remodeling, a model has emerged that maternal proteins might be sufficient for transforming tightly packaged sperm DNA into a fully competent paternal pronucleus (Levine, 2015).

The notion that maternal control is sufficient to accomplish paternal genome remodeling is challenged by recent findings from the intracellular Wolbachia bacterium that infects more than 50% of insect species. Wolbachia-infected Drosophila males mated to uninfected females father embryos that arrest soon after the first zygotic mitosis. Embryonic arrest occurs because paternal genomes enter the first mitosis with unresolved sister chromatids that fail to separate on the mitotic spindle. Although the identity of the host factor(s) manipulated by Wolbachia to mediate this transgenerational effect is still unknown, what is clear is that pre-fertilization, Wolbachia subverts the paternal germline machinery that helps direct global genome remodeling of paternal chromosomes in the embryo. Wolbachia action during spermiogenesis leads to paternal-maternal genome asynchrony and ultimately, failure of paternal chromosomes to separate on the first mitotic spindle. Despite decades of interest, the molecular basis of paternal control has remained elusive (Levine, 2015).

To investigate the potential for paternal control over sperm genome remodeling post-fertilization, a candidate gene approach was taken, focusing on the Heterochromatin Protein 1 (HP1) proteins that orchestrate genome-wide chromosomal organization in plants, animals, fungi, and some protists. HP1 proteins are defined as such by a combination of two domains - a chromodomain that mediates protein-histone interactions and a chromoshadow domain that mediates protein-protein interactions. The biochemical properties of HP1 members (Canzio, 2014) support a diversity of chromatin-dependent processes in the soma, including DNA replication, telomere integrity, and chromosome condensation (Levine, 2015).

Recently, a detailed phylogenomic analysis of the HP1 gene family was carried out in Drosophila that revealed numerous testis-restricted HP1 proteins (Levine, 2012). Given the established roles of HP1 proteins, it was posited that these newly discovered male-specific HP1 genes might represent excellent candidates for encoding chromatin functions specialized for paternal genome organization and remodeling in the early embryo. Using detailed genetic and cytological analyses, this study shows that one of these testis-specific HP1 proteins, Heterochromatin Protein 1E (HP1E), is essential for priming the paternal genome to enter embryonic mitosis in synchrony with the maternal genome in D. melanogaster. Intriguingly, HP1E is able to mediate this priming function transgenerationally i.e., the HP1E protein itself is not epigenetically inherited. It was further shown that absence of HP1E especially imperils mitotic fidelity of the heterochromatin-rich, paternal sex chromosomes. Thus, this study firmly establishes that both maternal and paternal control are necessary for paternal genome remodeling in the early Drosophila embryo (Levine, 2015).

Properly coordinated chromosome segregation during virtually all mitotic divisions relies on the function of multiple cell cycle checkpoint proteins. No such cell cycle checkpoint proteins have been identified to act in the very first embryonic mitotic cycle, which must nevertheless accomplish the difficult task of synchronizing maternal and paternal chromosomes that were inherited in very different chromatin states. To investigate the paternal contributions that ensure timely participation of the paternal genome in early embryogenesis, a detailed functional analysis was carried out of the testis-restricted HP1E gene in D. melanogaster. It was found that HP1E encodes a novel function that ensures paternal genome stability in the embryo. Cytological and transcriptome analysis revealed that HP1E is developmentally restricted within the male germline, where it contributes to heterochromatin integrity. HP1E depletion during sperm development results in a highly penetrant PEL phenotype in which paternal chromosomes, especially the paternal sex chromosomes, fail to condense in synchrony with the maternal chromosomes and ultimately cause mitotic catastrophe. It was further shown that the PEL embryonic phenotype could not be rescued by egg-supplied HP1E but could be rescued if the paternal DNA was excluded from participating in embryonic mitosis. These observations support a model under which HP1E acts pre-fertilization to ensure proper chromosome condensation and segregation of paternal chromosomes post-fertilization (Levine, 2015).

The 'hit and run' priming function clearly distinguishes HP1E from all other previously characterized paternal effect lethal genes, which encode proteins that are transmitted to the embryo via sperm. These include the Drosophila paternal chromatin-associated PEL, k81, which encodes a protein that persists on paternal telomeres from late spermatogenesis to the first embryonic mitosis. The HP1E-depletion phenotype is instead reminiscent of Drosophila fathers infected with Wolbachia bacteria crossed to uninfected females. Embryonic lethality induced by Wolbachia testis infection is also caused by a pre-fertilization modification to the paternal genome that results in paternal-maternal chromatin asynchrony and mis-segregation at the very first zygotic mitosis. However, Wolbachia-associated PEL results in mis-segregation of the entire paternal genome rather than just the heterochromatin-rich chromosomes observed in HP1E PEL . Moreover, the HP1E PEL defect is completely independent of Wolbachia (PEL phenotype persists for Wolbachia-free males and females). It is therefore concluded that HP1E supports a novel chromatin requirement to prime paternally inherited genomes for synchronous and successful embryonic mitosis (Levine, 2015).

How does HP1E ensure timely mitotic entry? It is formally possible that the PEL phenotype is the consequence of a dysregulated spermatid transcriptome that is, up- or down-regulation of a downstream gene. However, the finding that HP1E depletion results in the global up-regulation of heterochromatin-embedded genes, together with the observation that the heterochromatin-rich paternal sex chromosomes are most vulnerable to HP1E depletion, lead to favoring the alternate model that HP1E functions as a canonical HP1 protein during spermiogenesis. Based on antibody localization and chromatin bridge morphology, no evidence was found for defects in kinetochore assembly or replication machinery engagement in PEL embryos. Instead, the observation that the lethality phenotype first manifests as decondensed paternal chromosomes relative to maternal chromosomes implicates condensation delay of the heterochromatin-rich sex chromosomes. This delay could be the consequence of incomplete replication. Indeed, large stretches of uninterrupted heterochromatic DNA, as found on the Drosophila sex chromosomes, pose a unique challenge to replication. Alternatively, the mitotic delay may be the result of inadequate condensin protein recruitment, which is required for timely resolution of sister chromatids post-replication. Previous studies have shown that heterochromatin can also impair chromosome condensation. Timely completion of replication and condensation requires the action of HP1E's closest relative, HP1A, in somatic cells. However, in developing spermatids, HP1A localizes to telomeres rather than broadly to heterochromatin as observed in virtually all other cell types. It is posited that HP1E adopts a global, HP1A-like chromatin function during this highly specialized developmental stage and ensures the recruitment or retention of either replication or condensin proteins that are required post-fertilization (Levine, 2015).

Previous studies have shown that HP1A is essential for embryo viability. This study shows that paternally-acting HP1E is also essential for embryogenesis. Both HP1A and HP1E evolve under purifying selection. However, unlike HP1A (encoded by Su(var)205), HP1E has an unusually dynamic evolutionary history. Despite ancient origins, HP1E has been recurrently lost over evolutionary time. HP1E has been apparently replaced by younger, testis restricted HP1 paralogs on at least two occasions during Drosophila evolution (Levine, 2012). Curiously, Drosophila pseudoobscura and related species encode neither HP1E nor a putative replacement testis-specific HP1 gene. How is the paradox of HP1E essentiality in D. melanogaster reconciled with its loss in D. pseudoobscura? It was previously found that HP1E loss along in D. pseudoobscura-related species occurred during the same 7-million evolutionary period as a major sex chromosome rearrangement event (Levine, 2012), in which the ancestral Y was lost, a neo-Y chromosome was born, and the ancestral X fused to an autosome. The finding that the D. melanogaster sex chromosomes are especially vulnerable to HP1E depletion, combined with the emergence of novel sex chromosome arrangements along the same narrow branch as HP1E pseudogenization, suggests a model under which rearrangements of heterochromatin-rich sex chromosomes in the obscuragroup rendered HP1E non-essential. Such karyotypic changes can bring distal heterochromatin into closer proximity to euchromatin and be sufficient to alter heterochromatin packaging, replication timing or even delete blocks of satellite repeats. Thus, heterochromatin evolution via chromosomal rearrangements may have obviated maintenance of HP1E's essential heterochromatin function, leading to its degeneration in D. pseudoobscura (Levine, 2015).

The finding that HP1E is essential in D. melanogaster yet lost in the obscura group highlights the lineage-restricted essential requirements of chromatin genes. Intriguingly, the only other characterized PEL gene that supports paternal chromatin function in Drosophila embryos, k81, is similarly lineage-restricted despite being essential for paternal telomere function. In contrast, maternally deposited proteins required for paternal chromatin reorganization following fertilization are generally conserved from fly to human. This dichotomy is striking. It specifically suggests that even though the essential functions of paternal control of DNA deposition and chromatin remodeling for embryonic mitosis are likely to be conserved in most animals, whereas the identity of those genes is not. PEL chromatin genes like HP1E and k81 thus challenge the dogma that ancient, conserved genes always encode essential conserved functions. Not only can young genes rapidly acquire essential chromatin functions due to dynamic chromatin evolution, but chromatin changes, such as those driven by karyotype evolution, may also drive the extinction of ancient genes encoding once-essential functions (Levine, 2015).

Search PubMed for articles about Drosophila Hp1e

Canzio, D., Larson, A. and Narlikar, G. J. (2014). Mechanisms of functional promiscuity by HP1 proteins. Trends Cell Biol 24: 377-386. PubMed ID: 24618358

Delabaere, L., Orsi, G. A., Sapey-Triomphe, L., Horard, B., Couble, P. and Loppin, B. (2014). The Spartan ortholog maternal haploid is required for paternal chromosome integrity in the Drosophila zygote. Curr Biol 24: 2281-2287. PubMed ID: 25242033

Levine, M. T., McCoy, C., Vermaak, D., Lee, Y. C., Hiatt, M. A., Matsen, F. A. and Malik, H. S. (2012). Phylogenomic analysis reveals dynamic evolutionary history of the Drosophila heterochromatin protein 1 (HP1) gene family. PLoS Genet 8: e1002729. PubMed ID: 22737079

Levine, M.T., Vander Wende, H.M. and Malik, H.S. (2015). Mitotic fidelity requires transgenerational action of a testis-restricted HP1. Elife 4. PubMed ID: 26151671

McLay, D. W. and Clarke, H. J. (2003). Remodelling the paternal chromatin at fertilization in mammals. Reproduction 125: 625-633. PubMed ID: 12713425

Orsi, G. A., Algazeery, A., Meyer, R. E., Capri, M., Sapey-Triomphe, L. M., Horard, B., Gruffat, H., Couble, P., Ait-Ahmed, O. and Loppin, B. (2013). Drosophila Yemanuclein and HIRA cooperate for de novo assembly of H3.3-containing nucleosomes in the male pronucleus. PLoS Genet 9: e1003285. PubMed ID: 23408912

date revised: 3 August 2015

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