In vivo rescue of genetic dilated cardiomyopathy by systemic delivery of nexilin.

Background: Dilated cardiomyopathy (DCM) is one of the most common causes of heart failure. Multiple identified mutations in nexilin (NEXN) have been suggested to be linked with severe DCM. However, the exact association between multiple mutations of Nexn and DCM remains unclear.

Modulation of the microhomology-mediated end joining pathway suppresses large deletions and enhances homology-directed repair following CRISPR-Cas9-induced DNA breaks.

Background: CRISPR-Cas9 genome editing often induces unintended, large genomic rearrangements, posing potential safety risks. However, there are no methods for mitigating these risks.

Results: Using long-read individual-molecule sequencing (IDMseq), we found the microhomology-mediated end joining (MMEJ) DNA repair pathway plays a predominant role in Cas9-induced large deletions (LDs).

Quantitative haplotype-resolved analysis of mitochondrial DNA heteroplasmy in Human single oocytes, blastoids, and pluripotent stem cells.

Maternal mitochondria are the sole source of mtDNA for every cell of the offspring. Heteroplasmic mtDNA mutations inherited from the oocyte are a common cause of metabolic diseases and associated with late-onset diseases. However, the origin and dynamics of mtDNA heteroplasmy remain unclear.

Single-cell individual full-length mtDNA sequencing by iMiGseq uncovers unexpected heteroplasmy shifts in mtDNA editing.

The ontogeny and dynamics of mtDNA heteroplasmy remain unclear due to limitations of current mtDNA sequencing methods. We developed individual Mitochondrial Genome sequencing (iMiGseq) of full-length mtDNA for ultra-sensitive variant detection, complete haplotyping, and unbiased evaluation of heteroplasmy levels, all at the individual mtDNA molecule level. iMiGseq uncovered unappreciated levels of heteroplasmic variants in single cells well below the conventional NGS detection limit and provided accurate quantitation of heteroplasmy level.

Wiskott-Aldrich syndrome protein forms nuclear condensates and regulates alternative splicing.

The diverse functions of WASP, the deficiency of which causes Wiskott-Aldrich syndrome (WAS), remain poorly defined. We generated three isogenic WAS models using patient induced pluripotent stem cells and genome editing. These models recapitulated WAS phenotypes and revealed that WASP deficiency causes an upregulation of numerous RNA splicing factors and widespread altered splicing.

Structure of the full-length human Pannexin1 channel and insights into its role in pyroptosis.

Pannexin1 (PANX1) is a large-pore ATP efflux channel with a broad distribution, which allows the exchange of molecules and ions smaller than 1 kDa between the cytoplasm and extracellular space. In this study, we show that in human macrophages PANX1 expression is upregulated by diverse stimuli that promote pyroptosis, which is reminiscent of the previously reported lipopolysaccharide-induced upregulation of PANX1 during inflammasome activation. To further elucidate the function of PANX1, we propose the full-length human Pannexin1 (hPANX1) model through cryo-electron microscopy (cryo-EM) and molecular dynamics (MD) simulation studies, establishing hPANX1 as a homo-heptamer and revealing that both the N-termini and C-termini protrude deeply into the channel pore funnel.

Long-read individual-molecule sequencing reveals CRISPR-induced genetic heterogeneity in human ESCs.

Quantifying the genetic heterogeneity of a cell population is essential to understanding of biological systems. We develop a universal method to label individual DNA molecules for single-base-resolution haplotype-resolved quantitative characterization of diverse types of rare variants, with frequency as low as 4 × 10, using both short- or long-read sequencing platforms. It provides the first quantitative evidence of persistent nonrandom large structural variants and an increase in single-nucleotide variants at the on-target locus following repair of double-strand breaks induced by CRISPR-Cas9 in human embryonic stem cells.

The Kiss2/GPR54 system stimulates the reproductive axis in male black porgy, Acanthopagrus schlegelii.

Although it is well established that the Kiss1/GPR54 system stimulates the reproductive axis in mammals, its functional roles, especially in male reproduction of non-mammalian species, is less clear. In this study, we have isolated the full-length kiss2 and gpr54 cDNAs from black porgy (Acanthopagrus schlegelii). The Kiss2 precursor expressed from kiss2 comprises 124 amino acids and contains a highly conserved 10-amino acid sequence, Kiss2-10 (FNFNPFGLRF).

A 54-kDa short variant of DHX33 functions in regulating mRNA translation.

DEAH box protein DHX33 has been found to be necessary for cell proliferation and early development of multicellular organisms. It plays diverse roles in regulating gene transcription, ribosome RNA synthesis, and protein translation. Dysregulation of DHX33 has been observed in various human cancers.

Role of DHX33 in c-Myc-induced cancers.

Oncogene c-Myc is frequently amplified and activated in human cancers. Deregulation of c-Myc protein has been shown to occur in 30% of all human cancers, especially in hematopoietic malignancies. As a transcription factor, c-Myc has been shown to regulate up to 15% of all human genome genes, controlling diverse cellular activities including cell cycle, ribosome biogenesis, protein synthesis, metabolism, apoptosis and angiogenesis.

DDX59 promotes DNA replication in lung adenocarcinoma.

DEAD box proteins are multifunctional proteins involved in every aspect in RNA metabolism and have essential roles in many cellular activities. Despite their importance, many DEAD box proteins remain uncharacterized. In this report, we found DDX59 overexpressed in lung adenocarcinoma.

DHX33 Transcriptionally Controls Genes Involved in the Cell Cycle.

The RNA helicase DHX33 has been shown to be a critical regulator of cell proliferation and growth. However, the underlying mechanisms behind DHX33 function remain incompletely understood. We present original evidence in multiple cell lines that DHX33 transcriptionally controls the expression of genes involved in the cell cycle, notably cyclin, E2F1, cell division cycle (CDC), and minichromosome maintenance (MCM) genes.