CRISPR/Cas9 is a versatile genome-editing technology that is widely used for studying the functionality of genetic elements, creating genetically modified organisms as well as preclinical research of genetic disorders. However, the high frequency of off-target activity (≥50%)—RGEN (RNA-guided endonuclease)-induced mutations at sites other than the intended on-target site—is one major concern, especially for therapeutic and clinical applications. Here, we review the basic mechanisms underlying off-target cutting in the CRISPR/Cas9 system, methods for detecting off-target mutations, and strategies for minimizing off-target cleavage. The improvement off-target specificity in the CRISPR/Cas9 system will provide solid genotype–phenotype correlations, and thus enable faithful interpretation of genome-editing data, which will certainly facilitate the basic and clinical application of this technology.

Embryo implantation, a crucial step in human reproduction, is tightly controlled by estrogen and progesterone (P ₄ ) via estrogen receptor alpha and progesterone receptor (PGR), respectively. Here, we report that N ⁶ -methyladenosine (m ⁶ A), the most abundant mRNA modification in eukaryotes, plays an essential role in embryo implantation through the maintenance of P ₄ signaling. Conditional deletion of methyltransferase-like 3 ( Mettl3 ), encoding the m ⁶ A writer METTL3, in the female reproductive tract using a Cre mouse line with Pgr promoter ( Pgr-Cre ) resulted in complete implantation failure due to pre-implantation embryo loss and defective uterine receptivity. Moreover, the uterus of Mettl3 null mice failed to respond to artificial decidualization. We further found that Mettl3 deletion was accompanied by a marked decrease in PGR protein expression. Mechanistically, we found that Pgr mRNA is a direct target for METTL3-mediated m ⁶ A modification. A luciferase assay revealed that the m ⁶ A modification in the 5′ untranslated region (5′-UTR) of Pgr mRNA enhances PGR protein translation efficiency in a YTHDF1-dependent manner. Finally, we demonstrated that METTL3 is required for human endometrial stromal cell decidualization in vitro and that the METTL3-PGR axis is conserved between mice and humans. In summary, this study provides evidence that METTL3 is essential for normal P ₄ signaling during embryo implantation via m ⁶ A-mediated translation control of Pgr mRNA.

Dai is a major Chinese ethnic minority group residing in rural areas of the southern part of Yunnan. However, no data exist on the Human Papillomavirus (HPV) prevalence and genotype distribution among Dai women. A total of 793 participants (Dai=324, Han=251, other ethnic=218) were included in this study. PCR was performed to detect the HPV-positive samples, and genotyping was performed with an HPV Geno-Array. The overall HPV prevalence was very low among Dai women compared to the others. The prevalence of high-risk-HPV infections was significantly higher (p=0.001) among other ethnic women (22.0%) than that among Han (13.1%) and Dai women (7.1%). The overall HPV, high-risk-HPV, single and multiple infection prevalence among rural women were 12.9%, 12.1%, 12.3%, and 0.5%, respectively. HPV-16 (5.5%) was shown to be the most prevalent genotype, followed by HPV-52 (2.6%) and HPV-58 (2.4%). Urban women had relatively higher rates of overall HPV (16.0%), high-risk-HPV (14.1%), single genotype (11.9%), and multiple genotype (4.1%) infections. In urban women, HPV-52 (3.6%) was the most prevalent genotype, followed by HPV-39 (2.7%) and HPV-16 (1.2%). In the urban area, HPV prevalence was highest in women aged 50 years). Education and smoking habit were significantly associated with HPV infection among both rural and urban area women (p<0.001). The prevalence and genotype distribution of HPV varied among ethnic women in urban and rural area of Yunnan Province.

BACKGROUND:It is important to understand the knowledge that various groups of a population have about cervical cancer and human papillomavirus (HPV) and their attitudes toward HPV vaccination, as it will ultimately influence their decision-making for...

Gene editing in non-human primates may lead to valuable models for exploring the etiologies and therapeutic strategies of genetically based neurological disorders in humans. However, a monkey model of neurological disorders that closely mimics pathological and behavioral deficits in humans has not yet been successfully generated. Microcephalin 1 (MCPH1) is implicated in the evolution of the human brain, and MCPH1 mutation causes microcephaly accompanied by mental retardation. Here we generated a cynomolgus monkey (Macaca fascicularis) carrying biallelic MCPH1 mutations using transcription activator-like effector nucleases. The monkey recapitulated most of the important clinical features observed in patients, including marked reductions in head circumference, premature chromosome condensation (PCC), hypoplasia of the corpus callosum and upper limb spasticity. Moreover, overexpression of MCPH1 in mutated dermal fibroblasts rescued the PCC syndrome. This monkey model may help us elucidate the role of MCPH1 in the pathogenesis of human microcephaly and better understand the function of this protein in the evolution of primate brain size.