LOXL2-mediated H3K4 oxidation reduces chromatin accessibility in triple-negative breast cancer cells.

Oxidation of H3 at lysine 4 (H3K4ox) by lysyl oxidase-like 2 (LOXL2) generates an H3 modification with an unknown physiological function. We find that LOXL2 and H3K4ox are higher in triple-negative breast cancer (TNBC) cell lines and patient-derived xenografts (PDXs) than those from other breast cancer subtypes. ChIP-seq revealed that H3K4ox is located primarily in heterochromatin, where it is involved in chromatin compaction. Knocking down LOXL2 reduces H3K4ox levels and causes chromatin decompaction, resulting in a sustained activation of the DNA damage response (DDR) and increased susceptibility to anticancer agents. This critical role that LOXL2 and oxidized H3 play in chromatin compaction and DDR suggests that functionally targeting LOXL2 could be a way to sensitize TNBC cells to conventional therapy.

A genetic screen identifies a cellular regulator of adeno-associated virus.

Adeno-associated virus type 2 (AAV2) is a human parvovirus that has attracted attention as a vector for gene transfer. Replication and site-specific integration of the wild-type virus requires binding of the AAV2 Rep proteins to a cis-regulatory element named the Rep recognition sequence (RRS). RRS motifs are found within the cellular AAVS1 integration locus, the viral p5 promoter, and the inverted terminal repeats (ITRs). Here we report the design of a genetic screen based on the yeast one-hybrid assay to identify cellular RRS-binding proteins. We show that the human zinc finger 5 protein (ZF5) binds specifically to RRS motifs in vitro and in vivo. ZF5 is a highly conserved and ubiquitously expressed transcription factor that contains five C-terminal zinc fingers and an N-terminal POZ domain. Ectopic expression of ZF5 leads to an ITR-dependent repression of the autologous p5 promoter and reduces both AAV2 replication and the production of recombinant AAV2. By using deletion and substitution mutants we show that two different domains of ZF5 contribute to AAV2 repression. Negative regulation of the p5 promoter requires the POZ domain, whereas viral replication is inhibited by the zinc finger domain, likely by competing with Rep for binding to the ITR. Identification and characterization of proteins that bind the ITR, the only viral genetic element retained in AAV2 vectors, will lead to new insights into the unique life cycle of AAV2 and will suggest improvements important for its application as a gene therapy vector.

Identification of a steroidogenic neurohormone in female mosquitoes.

In the female mosquito, Aedes aegypti, neurohormones are released from the brain in response to a blood meal and stimulate the ovaries to secrete ecdysteroid hormones, which modulate yolk protein synthesis in the fat body. Neuropeptides with this bioactivity were isolated from head extracts, and partial sequences from these peptides when aligned gave a 31-residue sequence at the amino terminus. Oligonucleotide primers for this sequence were used to amplify with the polymerase chain reaction a genomic DNA product that hybridized to a clone from a head cDNA library. The cDNA encodes a 149-residue preprohormone that is processed into an 86-residue peptide, as indicated by the mass value obtained from the native peptide, with the expected amino-terminal sequence. After modification, the cDNA for the putative neurohormone was expressed in a bacterial system, and the purified peptide had high specific activity in bioassays, thus confirming that it is a steroidogenic gonadotropin, the first to be identified for invertebrates.