RESUMEN
Leukodystrophies are a heterogeneous group of rare inherited disorders that mostly involve the white matter of the CNS. These conditions are characterized by primary glial cell and myelin sheath pathology of variable aetiology, which causes secondary axonal degeneration, generally emerging with disease progression. Whole exome sequencing performed in five large consanguineous nuclear families allowed us to identify homozygosity for two recurrent missense variants affecting highly conserved residues of RNF220 as the causative event underlying a novel form of leukodystrophy with ataxia and sensorineural deafness. We report these two homozygous missense variants (p.R363Q and p.R365Q) in the ubiquitin E3 ligase RNF220 as the underlying cause of this novel form of leukodystrophy with ataxia and sensorineural deafness that includes fibrotic cardiomyopathy and hepatopathy as associated features in seven consanguineous families. Mass spectrometry analysis identified lamin B1 as the RNF220 binding protein and co-immunoprecipitation experiments demonstrated reduced binding of both RNF220 mutants to lamin B1. We demonstrate that RNF220 silencing in Drosophila melanogaster specifically affects proper localization of lamin Dm0, the fly lamin B1 orthologue, promotes its aggregation and causes a neurodegenerative phenotype, strongly supporting the functional link between RNF220 and lamin B1. Finally, we demonstrate that RNF220 plays a crucial role in the maintenance of nuclear morphology; mutations in primary skin fibroblasts determine nuclear abnormalities such as blebs, herniations and invaginations, which are typically observed in cells of patients affected by laminopathies. Overall, our data identify RNF220 as a gene implicated in leukodystrophy with ataxia and sensorineural deafness and document a critical role of RNF220 in the regulation of nuclear lamina. Our findings provide further evidence on the direct link between nuclear lamina dysfunction and neurodegeneration.
Asunto(s)
Alelos , Ataxia/genética , Sordera/genética , Laminopatías/genética , Mutación/genética , Ubiquitina-Proteína Ligasas/genética , Adolescente , Secuencia de Aminoácidos , Animales , Ataxia/diagnóstico , Células COS , Niño , Chlorocebus aethiops , Sordera/diagnóstico , Drosophila , Femenino , Células HEK293 , Humanos , Laminopatías/diagnóstico , Masculino , Linaje , Adulto JovenRESUMEN
Many genes are required for the assembly of the mitotic apparatus and for proper chromosome behavior during mitosis and meiosis. A fruitful approach to elucidate the mechanisms underlying cell division is the accurate phenotypic characterization of mutations in these genes. Here, we report the identification and characterization of diamond (dind), an essential Drosophila gene required both for mitosis of larval brain cells and for male meiosis. Larvae homozygous for any of the five EMS-induced mutations die in the third-instar stage and exhibit multiple mitotic defects. Mutant brain cells exhibit poorly condensed chromosomes and frequent chromosome breaks and rearrangements; they also show centriole fragmentation, disorganized mitotic spindles, defective chromosome segregation, endoreduplicated metaphases, and hyperploid and polyploid cells. Comparable phenotypes occur in mutant spermatogonia and spermatocytes. The dind gene encodes a non-conserved protein with no known functional motifs. Although the Dind protein exhibits a rather diffuse localization in both interphase and mitotic cells, fractionation experiments indicate that some Dind is tightly associated with the chromatin. Collectively, these results suggest that loss of Dind affects chromatin organization leading to defects in chromosome condensation and integrity, which in turn affect centriole stability and spindle assembly. However, our results do not exclude the possibility that Dind directly affects some behaviors of the spindle and centrosomes.
Asunto(s)
Cromosomas de Insectos/genética , Proteínas de Drosophila/genética , Drosophila/citología , Meiosis , Espermatocitos/fisiología , Animales , Animales Modificados Genéticamente , Encéfalo/citología , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , División Celular/genética , Rotura Cromosómica , Segregación Cromosómica , Drosophila/genética , Proteínas de Drosophila/metabolismo , Proteínas Fluorescentes Verdes/genética , Larva/citología , Masculino , Mutación , Fenotipo , Espermatocitos/citologíaRESUMEN
Drosophila telomeres are sequence-independent structures maintained by transposition to chromosome ends of three specialized retroelements rather than by telomerase activity. Fly telomeres are protected by the terminin complex that includes the HOAP, HipHop, Moi and Ver proteins. These are fast evolving, non-conserved proteins that localize and function exclusively at telomeres, protecting them from fusion events. We have previously suggested that terminin is the functional analogue of shelterin, the multi-protein complex that protects human telomeres. Here, we use electrophoretic mobility shift assay (EMSA) and atomic force microscopy (AFM) to show that Ver preferentially binds single-stranded DNA (ssDNA) with no sequence specificity. We also show that Moi and Ver form a complex in vivo. Although these two proteins are mutually dependent for their localization at telomeres, Moi neither binds ssDNA nor facilitates Ver binding to ssDNA. Consistent with these results, we found that Ver-depleted telomeres form RPA and γH2AX foci, like the human telomeres lacking the ssDNA-binding POT1 protein. Collectively, our findings suggest that Drosophila telomeres possess a ssDNA overhang like the other eukaryotes, and that the terminin complex is architecturally and functionally similar to shelterin.
Asunto(s)
Daño del ADN , ADN de Cadena Simple/metabolismo , Proteínas de Drosophila/metabolismo , Proteínas de Unión a Telómeros/metabolismo , Telómero/metabolismo , Animales , Proteínas Cromosómicas no Histona/fisiología , Reparación del ADN , ADN de Cadena Simple/ultraestructura , Drosophila/genética , Proteínas de Drosophila/química , Proteínas de Drosophila/fisiología , Proteínas de Drosophila/ultraestructura , Microscopía de Fuerza Atómica , Dominios Proteicos , Multimerización de Proteína , Proteína de Replicación A/metabolismo , Proteínas de Unión a Telómeros/química , Proteínas de Unión a Telómeros/ultraestructuraRESUMEN
Natural background radiation of Earth and cosmic rays played a relevant role during the evolution of living organisms. However, how chronic low doses of radiation can affect biological processes is still unclear. Previous data have indicated that cells grown at the Gran Sasso Underground Laboratory (LNGS, L'Aquila) of National Institute of Nuclear Physics (INFN) of Italy, where the dose rate of cosmic rays and neutrons is significantly reduced with respect to the external environment, elicited an impaired response against endogenous damage as compared to cells grown outside LNGS. This suggests that environmental radiation contributes to the development of defense mechanisms at cellular level. To further understand how environmental radiation affects metabolism of living organisms, we have recently launched the FLYINGLOW program that aims at exploiting Drosophila melanogaster as a model for evaluating the effects of low doses/dose rates of radiation at the organismal level. Here, we will present a comparative data set on lifespan, motility and fertility from different Drosophila strains grown in parallel at LNGS and in a reference laboratory at the University of L'Aquila. Our data suggest the reduced radiation environment can influence Drosophila development and, depending on the genetic background, may affect viability for several generations even when flies are moved back to normal background radiation. As flies are considered a valuable model for human biology, our results might shed some light on understanding the effect of low dose radiation also in humans.
Asunto(s)
Radiación de Fondo/efectos adversos , Drosophila melanogaster/efectos de la radiación , Fertilidad/efectos de la radiación , Longevidad/efectos de la radiación , Dosis de Radiación , Exposición a la Radiación/efectos adversos , Factores de Edad , Animales , Proteínas de la Ataxia Telangiectasia Mutada/genética , Conducta Animal/efectos de la radiación , Radiación Cósmica/efectos adversos , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Drosophila melanogaster/crecimiento & desarrollo , Femenino , Regulación del Desarrollo de la Expresión Génica/efectos de la radiación , Genotipo , Locomoción/efectos de la radiación , Masculino , Mutación , Neutrones/efectos adversos , Fenotipo , Proteínas Serina-Treonina QuinasasRESUMEN
Multicolor immunostaining analysis is often a desirable tool in cell biology for most researchers. Nonetheless, this is not an easy task and often not affordable by many laboratories as it might require expensive instrumentation and sophisticated analysis software. Here, we describe a simple protocol for performing sequential immunostainings on two different Drosophila specimens. Our strategy relies on an efficient and reproducible method for removal primary antibodies and/or fluorophore-conjugated secondary antibodies that does not affect antigene integrity. We show that alternation of multiple rounds of antibody incubation and removal on the same slide, followed by registration of the same DAPI-stained image, provides a simple framework for the sequential detection of several antigens in the same cell. Given that the sample fixation procedures used for Drosophila tissues are compatible with most specimen processing protocols, we can envisage that the multicolor immunostaining strategy presented here can be also adapted to different samples including mammalian tissues and/or cells.
Asunto(s)
Antígenos/análisis , Drosophila/citología , Técnica del Anticuerpo Fluorescente/métodos , Coloración y Etiquetado/métodos , Animales , Anticuerpos/química , Anticuerpos/inmunología , Cromatina/química , Cromatina/inmunología , Homólogo de la Proteína Chromobox 5 , Proteínas Cromosómicas no Histona/análisis , Color , Proteínas de Drosophila/análisis , Colorantes Fluorescentes , Histonas/análisis , Indoles , Larva/citología , Masculino , Cromosomas Politénicos/química , Cromosomas Politénicos/ultraestructura , Glándulas Salivales/citología , Espermatocitos/citología , Testículo/citología , Enzimas Ubiquitina-Conjugadoras/análisisRESUMEN
Telomeres are specialized structures at the end of eukaryotic chromosomes that are required to preserve genome integrity, chromosome stability and nuclear architecture. Telomere maintenance and function are established epigenetically in several eukaryotes. However, the exact chromatin enzymatic modifications regulating telomere homeostasis are poorly understood. In Drosophila melanogaster, telomere length and stability are maintained through the retrotransposition of specialized telomeric sequences and by the specific loading of protecting capping proteins, respectively. Here, we show that the loss of the essential and evolutionarily conserved histone deacetylase Rpd3, the homolog of mammalian HDAC1, causes aberrant telomeric fusions on polytene chromosome ends. Remarkably, these telomere fusion defects are associated with a marked decrease of histone H4 acetylation, as well as an accumulation of heterochromatic epigenetic marks at telomeres, including histone H3K9 trimethylation and the heterochromatic protein HP2. Our work suggests that Drosophila telomere structure is epigenetically regulated by the histone deacetylase Rpd3.
Asunto(s)
Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Heterocromatina/metabolismo , Histona Desacetilasa 1/metabolismo , Telómero/metabolismo , Animales , Drosophila melanogaster/metabolismo , Epigenómica , Masculino , Cromosomas PoliténicosRESUMEN
Chromobox Protein 3 (CBX3) overexpression is a common event occurring in cancer, promotes cancer cell proliferation and represents a poor prognosis marker in a plethora of human cancers. Here we describe that a wide spectrum of human cancers harbors a co-amplification of CBX3 gene with either EGFR or RAC1, which yields a statistically significant increase of both mRNA and protein levels of CBX3, EGFR and RAC1. We also reveal that the simultaneous overexpression of CBX3, RAC1 and EGFR gene products correlates with a worse prognosis compared to the condition when CBX3, RAC1 and EGFR are singularly upregulated. Furthermore, we also show that a co-occurrence of low-grade amplification, in addition to high-grade amplification, between CBX3 and EGFR or RAC1 is associated with a reduced patient lifespan. Finally, we find that CBX3 and RAC1/EGFR genetically interact in the model organism Drosophila melanogaster, suggesting that the simultaneous overexpression as well as well the co-occurrence of high- or low-grade copy number alterations in these genes is not accidental and could reflect evolutionarily conserved functional relationships.
RESUMEN
Compelling evidence indicates that defects in nucleocytoplasmic transport contribute to the pathogenesis of amyotrophic lateral sclerosis (ALS). In particular, hexanucleotide (G4C2) repeat expansions in C9orf72, the most common cause of genetic ALS, have a widespread impact on the transport machinery that regulates the nucleocytoplasmic distribution of proteins and RNAs. We previously reported that the expression of G4C2 hexanucleotide repeats in cultured human and mouse cells caused a marked accumulation of poly(A) mRNAs in the cell nuclei. To further characterize the process, we set out to systematically identify the specific mRNAs that are altered in their nucleocytoplasmic distribution in the presence of C9orf72-ALS RNA repeats. Interestingly, pathway analysis showed that the mRNAs involved in membrane trafficking are particularly enriched among the identified mRNAs. Most importantly, functional studies in cultured cells and Drosophila indicated that C9orf72 toxic species affect the membrane trafficking route regulated by ADP-Ribosylation Factor 1 GTPase Activating Protein (ArfGAP-1), which exerts its GTPase-activating function on the small GTPase ADP-ribosylation factor 1 to dissociate coat proteins from Golgi-derived vesicles. We demonstrate that the function of ArfGAP-1 is specifically affected by expanded C9orf72 RNA repeats, as well as by C9orf72-related dipeptide repeat proteins (C9-DPRs), indicating the retrograde Golgi-to-ER vesicle-mediated transport as a target of C9orf72 toxicity.
Asunto(s)
Esclerosis Amiotrófica Lateral , Proteína C9orf72 , Proteínas Activadoras de GTPasa , Animales , Humanos , Ratones , Factor 1 de Ribosilacion-ADP/metabolismo , Esclerosis Amiotrófica Lateral/metabolismo , Proteína C9orf72/genética , Proteína C9orf72/metabolismo , Drosophila/genética , Drosophila/metabolismo , ARN/metabolismo , ARN Mensajero/genética , Proteínas Activadoras de GTPasa/genética , Proteínas Activadoras de GTPasa/metabolismoRESUMEN
Telomeres in Drosophila melanogaster, which have inspired a large part of Sergio Pimpinelli work, are similar to those of other eukaryotes in terms of their function. Yet, their length maintenance relies on the transposition of the specialized retrotransposons Het-A, TART, and TAHRE, rather than on the activity of the enzyme telomerase as it occurs in most other eukaryotic organisms. The length of the telomeres in Drosophila thus depends on the number of copies of these transposable elements. Our previous work has led to the isolation of a dominant mutation, Tel1, that caused a several-fold elongation of telomeres. In this study, we molecularly identified the Tel1 mutation by a combination of transposon-induced, site-specific recombination and next-generation sequencing. Recombination located Tel1 to a 15 kb region in 92A. Comparison of the DNA sequence in this region with the Drosophila Genetic Reference Panel of wild-type genomic sequences delimited Tel1 to a 3 bp deletion inside intron 8 of Ino80. Furthermore, CRISPR/Cas9-induced deletions surrounding the same region exhibited the Tel1 telomere phenotype, confirming a strict requirement of this intron 8 gene sequence for a proper regulation of Drosophila telomere length.
Asunto(s)
Drosophila melanogaster , Drosophila , Animales , Drosophila/genética , Drosophila melanogaster/genética , Productos del Gen gag/genética , Telómero/genética , Mutación/genéticaRESUMEN
The Drosophila melanogasterDmATPCL gene encodes for the human ATP Citrate Lyase (ACL) ortholog, a metabolic enzyme that from citrate generates glucose-derived Acetyl-CoA, which fuels central biochemical reactions such as the synthesis of fatty acids, cholesterol and acetylcholine, and the acetylation of proteins and histones. We had previously reported that, although loss of Drosophila ATPCL reduced levels of Acetyl-CoA, unlike its human counterpart, it does not affect global histone acetylation and gene expression, suggesting that its role in histone acetylation is either partially redundant in Drosophila or compensated by alternative pathways. Here, we describe that depletion of DmATPCL affects spindle organization, cytokinesis, and fusome assembly during male meiosis, revealing an unanticipated role for DmATPCL during spermatogenesis. We also show that DmATPCL mutant meiotic phenotype is in part caused by a reduction of fatty acids, but not of triglycerides or cholesterol, indicating that DmATPCL-derived Acetyl-CoA is predominantly devoted to the biosynthesis of fatty acids during spermatogenesis. Collectively, our results unveil for the first time an involvement for DmATPCL in the regulation of meiotic cell division, which is likely conserved in human cells.
Asunto(s)
División Celular , Drosophila melanogaster/enzimología , Complejos Multienzimáticos/metabolismo , Oxo-Ácido-Liasas/metabolismo , Espermatogénesis , Animales , División Celular/genética , Masculino , Complejos Multienzimáticos/genética , Oxo-Ácido-Liasas/genética , Espermatogénesis/genéticaRESUMEN
Scientific community and institutions (e. g., ICRP) consider that the Linear No-Threshold (LNT) model, which extrapolates stochastic risk at low dose/low dose rate from the risk at moderate/high doses, provides a prudent basis for practical purposes of radiological protection. However, biological low dose/dose rate responses that challenge the LNT model have been highlighted and important dowels came from radiobiology studies conducted in Deep Underground Laboratories (DULs). These extreme ultra-low radiation environments are ideal locations to conduct below-background radiobiology experiments, interesting from basic and applied science. The INFN Gran Sasso National Laboratory (LNGS) (Italy) is the site where most of the underground radiobiological data has been collected so far and where the first in vivo underground experiment was carried out using Drosophila melanogaster as model organism. Presently, many DULs around the world have implemented dedicated programs, meetings and proposals. The general message coming from studies conducted in DULs using protozoan, bacteria, mammalian cells and organisms (flies, worms, fishes) is that environmental radiation may trigger biological mechanisms that can increase the capability to cope against stress. However, several issues are still open, among them: the role of the quality of the radiation spectrum in modulating the biological response, the dependence on the biological endpoint and on the model system considered, the overall effect at organism level (detrimental or beneficial). At LNGS, we recently launched the RENOIR experiment aimed at improving knowledge on the environmental radiation spectrum and to investigate the specific role of the gamma component on the biological response of Drosophila melanogaster.
Asunto(s)
Laboratorios , Protección Radiológica , Animales , Drosophila melanogaster , Italia , RadiobiologíaRESUMEN
Heterochromatin Protein 1 (HP1) and the Mre11-Rad50-Nbs1 (MRN) complex are conserved factors that play crucial role in genome stability and integrity. Despite their involvement in overlapping cellular functions, ranging from chromatin organization, telomere maintenance to DNA replication and repair, a tight functional relationship between HP1 and the MRN complex has never been elucidated. Here we show that the Drosophila HP1a protein binds to the MRN complex through its chromoshadow domain (CSD). In addition, loss of any of the MRN members reduces HP1a levels indicating that the MRN complex acts as regulator of HP1a stability. Moreover, overexpression of HP1a in nbs (but not in rad50 or mre11) mutant cells drastically reduces DNA damage associated with the loss of Nbs suggesting that HP1a and Nbs work in concert to maintain chromosome integrity in flies. We have also found that human HP1α and NBS1 interact with each other and that, similarly to Drosophila, siRNA-mediated inhibition of NBS1 reduces HP1α levels in human cultured cells. Surprisingly, fibroblasts from Nijmegen Breakage Syndrome (NBS) patients, carrying the 657del5 hypomorphic mutation in NBS1 and expressing the p26 and p70 NBS1 fragments, accumulate HP1α indicating that, differently from NBS1 knockout cells, the presence of truncated NBS1 extends HP1α turnover and/or promotes its stability. Remarkably, an siRNA-mediated reduction of HP1α in NBS fibroblasts decreases the hypersensitivity to irradiation, a characteristic of the NBS syndrome. Overall, our data provide an unanticipated evidence of a close interaction between HP1 and NBS1 that is essential for genome stability and point up HP1α as a potential target to counteract chromosome instability in NBS patient cells.
Asunto(s)
Proteínas de Ciclo Celular/genética , Proteínas Cromosómicas no Histona/genética , Proteínas de Drosophila/genética , Endodesoxirribonucleasas/genética , Exodesoxirribonucleasas/genética , Inestabilidad Genómica/genética , Proteínas Nucleares/genética , Animales , Homólogo de la Proteína Chromobox 5 , Daño del ADN/genética , Drosophila melanogaster/genética , Femenino , Fibroblastos/metabolismo , Fibroblastos/patología , Regulación de la Expresión Génica , Genoma de los Insectos/genética , Humanos , Masculino , Mutación/genética , Síndrome de Nijmegen/genética , Síndrome de Nijmegen/patologíaRESUMEN
Deep underground laboratories (DULs) were originally created to host particle, astroparticle or nuclear physics experiments requiring a low-background environment with vastly reduced levels of cosmic-ray particle interference. More recently, the range of science projects requiring an underground experiment site has greatly expanded, thus leading to the recognition of DULs as truly multidisciplinary science sites that host important studies in several fields, including geology, geophysics, climate and environmental sciences, technology/instrumentation development and biology. So far, underground biology experiments are ongoing or planned in a few of the currently operating DULs. Among these DULs is the Gran Sasso National Laboratory (LNGS), where the majority of radiobiological data have been collected. Here we provide a summary of the current scenario of DULs around the world, as well as the specific features of the LNGS and a summary of the results we obtained so far, together with other findings collected in different underground laboratories. In particular, we focus on the recent results from our studies of Drosophila melanogaster, which provide the first evidence of the influence of the radiation environment on life span, fertility and response to genotoxic stress at the organism level. Given the increasing interest in this field and the establishment of new projects, it is possible that in the near future more DULs will serve as sites of radiobiology experiments, thus providing further relevant biological information at extremely low-dose-rate radiation. Underground experiments can be nicely complemented with above-ground studies at increasing dose rate. A systematic study performed in different exposure scenarios provides a potential opportunity to address important radiation protection questions, such as the dose/dose-rate relationship for cancer and non-cancer risk, the possible existence of dose/dose-rate threshold(s) for different biological systems and/or end points and the possible role of radiation quality in triggering the biological response.
Asunto(s)
Radiación de Fondo , Daño del ADN/genética , Drosophila melanogaster/genética , Radiobiología/tendencias , Animales , Daño del ADN/efectos de la radiación , Relación Dosis-Respuesta en la Radiación , Drosophila melanogaster/efectos de la radiación , Protección RadiológicaRESUMEN
Drosophila telomeres are elongated by transposition of specialized retroelements rather than telomerase activity and are assembled independently of the sequence. Fly telomeres are protected by the terminin complex that localizes and functions exclusively at telomeres and by non-terminin proteins that do not serve telomere-specific functions. We show that mutations in the Drosophila Separase encoding gene Sse lead not only to endoreduplication but also telomeric fusions (TFs), suggesting a role for Sse in telomere capping. We demonstrate that Separase binds terminin proteins and HP1, and that it is enriched at telomeres. Furthermore, we show that loss of Sse strongly reduces HP1 levels, and that HP1 overexpression in Sse mutants suppresses TFs, suggesting that TFs are caused by a HP1 diminution. Finally, we find that siRNA-induced depletion of ESPL1, the Sse human orthologue, causes telomere dysfunction and HP1 level reduction in primary fibroblasts, highlighting a conserved role of Separase in telomere protection.
Asunto(s)
Proteínas de Drosophila/metabolismo , Separasa/metabolismo , Telómero/metabolismo , Animales , Proteínas Cromosómicas no Histona/genética , Proteínas Cromosómicas no Histona/metabolismo , Drosophila , Proteínas de Drosophila/genética , Humanos , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Separasa/genética , Telómero/genéticaRESUMEN
The Drosophila effete gene encodes an extremely conserved class I E2 ubiquitin-conjugating enzyme. Growing evidence indicates that Eff is involved in many cellular processes including eye development, maintenance of female germline stem cells, and regulation of apoptosis. Eff is also a major component of Drosophila chromatin and it is particularly enriched in chromatin with repressive properties. In addition, Eff is required for telomere protection and to prevent telomere fusion. Consistent with its multiple roles in chromatin maintenance, Eff is also one of the rare factors that modulate both telomere-induced and heterochromatin-induced position effect variegation.
Asunto(s)
Ensamble y Desensamble de Cromatina , Cromatina/metabolismo , Proteínas de Drosophila/fisiología , Drosophila/genética , Enzimas Ubiquitina-Conjugadoras/fisiología , Animales , Apoptosis/genética , Drosophila/crecimiento & desarrollo , Drosophila/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Femenino , Masculino , Homeostasis del Telómero/genética , Enzimas Ubiquitina-Conjugadoras/genética , Enzimas Ubiquitina-Conjugadoras/metabolismoRESUMEN
Drosophila telomeres are elongated by the transposition of telomere-specific retrotransposons rather than telomerase activity. Proximal to the terminal transposon array, Drosophila chromosomes contain several kilobases of a complex satellite DNA termed telomere-associated sequences (TASs). Reporter genes inserted into or next to the TAS are silenced through a mechanism called telomere position effect (TPE). TPE is reminiscent of the position effect variegation (PEV) induced by Drosophila constitutive heterochromatin. However, most genes that modulate PEV have no effect on TPE, and systematic searches for TPE modifiers have so far identified only a few dominant suppressors. Surprisingly, only a few of the genes required to prevent telomere fusion have been tested for their effect on TPE. Here, we show that with the exception of the effete (eff; also called UbcD1) mutant alleles, none of the tested mutations at the other telomere fusion genes affects TPE. We also found that mutations in eff, which encodes a class I ubiquitin-conjugating enzyme, act as suppressors of PEV. Thus, eff is one of the rare genes that can modulate both TPE and PEV. Immunolocalization experiments showed that Eff is a major constituent of polytene chromosomes. Eff is enriched at several euchromatic bands and interbands, the TAS regions, and the chromocenter. Our results suggest that Eff associates with different types of chromatin affecting their abilities to regulate gene expression.