The Drosophila Helicase Maleless (MLE) is Implicated in Functions Distinct From its Role in Dosage Compensation*

Abstract

Helicases are ubiquitous enzymes that unwind or remodel single or double-stranded nucleic acids, and that participate in a vast array of metabolic pathways. The ATP-dependent DEXH-box RNA/DNA helicase MLE was first identified as a core member of the chromatin remodeling MSL complex, responsible for dosage compensation in Drosophila males. Although this complex does not assemble in females, MLE is present. Given the multiplicity of functions attributed to its mammalian ortholog RNA helicase A, we have carried out an analysis for the purpose of determining whether MLE displays the same diversity. We have identified a number of different proteins that associate with MLE, implicating its role in specific pathways. We have documented this association in selected examples that include the spliceosome complex, Helicases are involved in all cellular transactions related to nucleic acid function and metabolism in prokaryotes and eukaryotes. These enzymes participate in transcription, RNA splicing, the stability of transcripts, and translation initiation, as well as in DNA-replication, repair, and recombination. Helicases use the energy produced by the hydrolysis of nucleotide triphosphates to catalyze the unwinding of DNA, RNA, and RNA:DNA hybrid molecules. They are also involved in protein displacement from RNA, RNA clamping, strand annealing, and RNA structure conversion (1). Eukaryotic helicases can be divided into two superfamilies, SF1 and SF2, which include three and nine families, respectively. Members of the different families can be distinguished on the basis of whether they use the hydrolysis of ATP or some other nucleotide triphosphates for energy release, whether they unwind DNA or RNA, and whether they require a 3Ј overhang or a 5Ј overhang to carry out their unwinding functions (2, 3).The Drosophila helicase maleless (MLE) 1 is a subunit of the MSL (male-specific lethal) complex that is responsible for dosage compensation -the regulatory mechanism involved in equalizing the levels of X chromosome-linked gene products between the sexes. In addition to MLE, the MSL complex contains an ubiquitin ligase (MSL2), a histone acetyl transferase (MOF), two structural proteins (MSL1 and MSL3), and one of two long non-coding RNAs (roX1 and roX2) that are necessary for its assembly and targeting. MLE is an ATP-dependent DEXH-box RNA/DNA helicase that in vitro prefers hybrid RNA:DNA or double-stranded RNA substrates with a short 3Ј overhang (4). MLE exhibits some similarity to the ATPases present in complexes that remodel chromatin by altering the positioning or the architectural relationship between histone octamers and DNA. In contrast to MLE, none of these enzymatic subunits have been shown to possess double-stranded nucleic acid unwinding activity. The absence or loss of function of MLE leads to failures in assembly and targeting of the MSL complex to its numerous sites of action along the X chromosome in males, although MSL1 and MSL2 are present at a few "high affinity" or "entry" sites (5-10). Recent biochemical ev...

The essential Drosophila CLAMP protein differentially regulates non-coding roX RNAs in male and females

“…We did not detect ectopic localization of MSL2 on either the X-chromosome or autosomes in clamp 2 heterozygous or homozygous females (Figure 4B, w − ; clamp 2 / CyO -GFP and w − ; clamp 2 / clamp 2 ). Furthermore, we detected MLE in similar non-X-specific patterns on clamp 2 mutant and wild type female polytene chromosomes, consistent with its known localization pattern (Figure 4B, y − w − and w − ; clamp 2 / clamp 2 ) (Cugusi et al , 2015). Therefore, the large increases in roX transcripts observed in clamp 2 homozygous females are not likely to be due to MSL complex-mediated roX activation.…”

Drosophila CLAMP is an essential protein with sex-specific roles in males and females

“…Therefore, if MSL complex forms in clamp 2 homozygous females, it is likely that it would not be capable of localizing to the X-chromosome. Additionally, we see minimal differences in localization of MLE, a non-sex specific complex component known to be expressed in females and to have functions distinct from its role in dosage compensation ( Figure 5B) (Cugusi et al, 2015).…”

The Drosophila Helicase MLE Targets Hairpin Structures in Genomic Transcripts

“…MLE’s RNA unwinding activity is also essential for its function in dosage compensation [ 14 ], where it is required for remodeling the RNA on the X (roX) RNAs’ loop structures in order to facilitate the assembly of the MSL complex [ 15 , 16 ]. Recently, we have obtained evidence that MLE is involved in a large number of additional regulatory steps and pathways involved in nucleic acid metabolism [ 17 ].…”