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 approximately 35 chromatin entry sites and subsequent spreading of mature complexes in cis to numerous additional sites along the chromosome. To understand this process further we examined MSL patterns in a range of wild-type and mutant backgrounds producing different amounts of MSL components. Our 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. We also mapped a set of definite autosomal regions (approximately 70) competent to associate with the functional MSL complex in wild-type males. 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.
In Drosophila, the dosage compensation complex (DCC) mediates upregulation of transcription from the single male X chromosome. Despite coating the polytene male X, the DCC pattern looks discontinuous and probably reflects DCC dynamic associations with genes active at a given moment of development in a salivary gland. To test this hypothesis, we compared binding patterns of the DCC and of the elongating form of RNA polymerase II (PolIIo). We found that, unlike PolIIo, the DCC demonstrates a stable banded pattern throughout larval development and escapes binding to a subset of transcriptionally active areas, including developmental puffs. Moreover, these proteins are not completely colocalized at the electron microscopy level. These data combined imply that simple recognition of PolII machinery or of general features of active chromatin is either insufficient or not involved in DCC recruitment to its targets. We propose that DCC-mediated site-specific upregulation of transcription is not the fate of all active X-linked genes in males. Additionally, we found that DCC subunit MLE associates dynamically with developmental and heat-shock-induced puffs and, surprisingly, with those developing within DCC-devoid regions of the male X, thus resembling the PolIIo pattern. These data imply that, independently of other MSL proteins, the RNA-helicase MLE might participate in general transcriptional regulation or RNA processing.
The morphological characteristics of intercalary heterochromatin (IH) are compared with those of other types of silenced chromatin in the Drosophila melanogaster genome: pericentric heterochromatin (PH) and regions subject to position effect variegation (PEV). We conclude that IH regions in polytene chromosomes are binding sites of silencing complexes such as PcG complexes and of SuUR protein. Binding of these proteins results in the appearance of condensed chromatin and late replication of DNA, which in turn may result in DNA underreplication. IH and PH as well as regions subject to PEV have in common the condensed chromatin appearance, the localization of specific proteins, late replication, underreplication in polytene chromosomes, and ectopic pairing.
Regions of intercalary heterochromatin (IH) are dispersed in the euchromatic arms of polytene chromosomes and share the main properties of heterochromatin, namely chromosome constrictions resulting from DNA underreplication. These constrictions are frequent on the paired X chromosomes of females, but are practically absent from the single X chromosome of males. These sex-specific differences have been proposed to reflect the different levels of transcription and chromosome compaction due to dosage compensation, which in turn may affect the degree of underreplication in IH regions. To test this hypothesis, we induced dosage compensation in females by ectopic expression of MSL-2 protein. We then measured the extent of underreplication in IH regions by determining frequencies of constrictions, or by Southern blot analysis using a fragment of the ten (a) gene which is located in IH region 11A6-9. Females transheterozygous for Sxl (fhv1)/ Sxl (f1) or carrying a constitutive msl-2 transgene are known to hypertranscribe their X chromosomes. In such females, both the frequency of constrictions and DNA underreplication were reduced. Suppression of underreplication occurs only when a complete functional MSL complex assembles on the X chromosomes. We also used three strains that carried constitutive transgenes of msl-2 with mutations in the 5' untranslated regions. These strains produced normal levels of SXL protein, but variable levels of MSL-2 protein. The SXL protein did not prevent the formation of an MSL complex in these transgenic females. We found that the extent of underreplication of ten (a) DNA in IH region 11A6-9 negatively correlates with the amount of MSL complex.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.