Risk assessment of airborne agricultural pesticide exposure in humans in rural China.

Continuous exposure to airborne pesticides causes their gradual accumulation in the human body, eventually posing a threat to human health. To the best of our knowledge, risk assessment study of pesticide non-occupational exposure to residents in agricultural areas has not been conducted in China. In this study, air samples (gas and dust) were collected from inside and outside residences of seven households and an area near the field in a grain-growing area (wheat and maize rotation) for eight months, and the pesticides present were examined both qualitatively and quantitatively. Using a 95% confidence interval, 9 out of 16 pesticides were detected, namely acetamiprid, acetochlor, atrazine, flucarbazone-sodium, imidacloprid, methyldisulfuron-methyl, nicosulfuron-methyl, pendimethalin, and beta-cyhalothrin, and their safety was subsequently evaluated. The results showed that the inhalation exposure of households to beta-cyhalothrin exceeded the acceptable range in the first residential, and the excess lifetime cancer risk of acetochlor inhalation exposure in six households and area around the field exceeds 1E-6, which highlights the need to strengthen preventive screening for cancer risk.

Risk assessment of human exposure to airborne pesticides in rural greenhouses.

In comparison to an open field, greenhouses utilize much more pesticides. The non-occupational exposure risk caused by pesticide drift is unknown. In this study, within 8 months (from March 2018 to October 2018), air samples were collected from indoor and outdoor houses and public areas near greenhouses in vegetable growing areas (eggplant, leek, garlic, etc.), and qualitative and quantitative analyses of pesticides were carried out. Using a 95% confidence interval, six pesticides (acetamiprid, difenoconazole, thiazophos, isoprocarb, malathion, and pyridaben) were detected. The results of the safety assessment showed that the non-cancer exposure risk of single pesticides for all residents in the agricultural areas was within the acceptable range, and the excess lifetime cancer risk of all residents inhaling difenoconazole exceeded 1E-6, and the agricultural region urgently needs increased cancer regulatory scrutiny. But combined toxicity of six pesticides not evaluated due to lack of suitable data. Comparison with open field scenes, the results show that pesticide levels to airborne are lower in greenhouse regions.

Cryptochromes and UBP12/13 deubiquitinases antagonistically regulate DNA damage response in Arabidopsis.

Cryptochromes (CRYs) are evolutionarily conserved blue-light receptors that evolved from bacterial photolyases that repair damaged DNA. Today, CRYs have lost their ability to repair damaged DNA; however, prior reports suggest that human CRYs can respond to DNA damage. Currently, the role of CRYs in the DNA damage response (DDR) is lacking, especially in plants. Therefore, we evaluated the role of plant CRYs in DDR along with UBP12/13 deubiquitinases, which interact with and regulate the CRY2 protein. We found that cry1cry2 was hypersensitive, while ubp12ubp13 was hyposensitive to UVC-induced DNA damage. Elevated UV-induced cyclobutane pyrimidine dimers (CPDs) and the lack of DNA repair protein RAD51 accumulation in cry1cry2 plants indicate that CRYs are required for DNA repair. On the contrary, CPD levels diminished and RAD51 protein levels elevated in plants lacking UBP12 and UBP13, indicating their role in DDR repression. Temporal transcriptomic analysis revealed that DDR-induced transcriptional responses were subdued in cry1cry2, but elevated in ubp12ubp13 compared to WT. Through transcriptional modeling of the time-course transcriptome, we found that genes quickly induced by UVC (15 min) are targets of CAMTA 1-3 transcription factors, which we found are required for DDR. This transcriptional regulation seems, however, diminished in the cry1cry2 mutant, indicating that CAMTAs are required for CRY2-mediated DDR. Furthermore, we observed enhanced CRY2-UBP13 interaction and formation of CRY2 nuclear speckles under UVC, suggesting that UVC activates CRY2 similarly to blue light. Together, our data reveal the temporal dynamics of the transcriptional events underlying UVC-induced genotoxicity and expand our knowledge of the role of CRY and UBP12/13 in DDR.

Systematic histone H4 replacement in Arabidopsis thaliana reveals a role for H4R17 in regulating flowering time.

Despite the broad array of roles for epigenetic mechanisms on regulating diverse processes in eukaryotes, no experimental system is currently available in plants for the direct assessment of histone function. In this work, we present the development of a genetic strategy in Arabidopsis (Arabidopsis thaliana) whereby modified histone H4 transgenes can completely replace the expression of endogenous histone H4 genes. Accordingly, we established a collection of plants expressing different H4 point mutants targeting residues that may be post-translationally modified in vivo. To demonstrate its utility, we screened this new H4 mutant collection to uncover substitutions in H4 that alter flowering time. We identified different mutations in the H4 tail (H4R17A) and the H4 globular domain (H4R36A, H4R39K, H4R39A, and H4K44A) that strongly accelerate the floral transition. Furthermore, we identified a conserved regulatory relationship between H4R17 and the ISWI chromatin remodeling complex in plants: As with other biological systems, H4R17 regulates nucleosome spacing via ISWI. Overall, this work provides a large set of H4 mutants to the plant epigenetics community that can be used to systematically assess histone H4 function in Arabidopsis and a roadmap to replicate this strategy for studying other histone proteins in plants.

UBP12 and UBP13 deubiquitinases destabilize the CRY2 blue light receptor to regulate Arabidopsis growth.

Light is a crucial exogenous signal sensed by cryptochrome (CRY) blue light receptors to modulate growth and the circadian clock in plants and animals. However, how CRYs interpret light quantity to regulate growth in plants remains poorly understood. Furthermore, CRY2 protein levels and activity are tightly regulated in light to fine-tune hypocotyl growth; however, details of the mechanisms that explain precise control of CRY2 levels are not fully understood. We show that in Arabidopsis, UBP12 and UBP13 deubiquitinases physically interact with CRY2 in light. UBP12/13 negatively regulates CRY2 by promoting its ubiquitination and turnover to modulate hypocotyl growth. Growth and development were explicitly affected in blue light when UBP12/13 were disrupted or overexpressed, indicating their role alongside CRY2. UBP12/13 also interacted with and stabilized COP1, which is partially required for CRY2 turnover. Our combined genetic and molecular data support a mechanistic model in which UBP12/13 interact with CRY2 and COP1, leading to the stabilization of COP1. Stabilized COP1 then promotes the ubiquitination and degradation of CRY2 under blue light. Despite decades of studies on deubiquitinases, the knowledge of how their activity is regulated is limited. Our study provides insight into how exogenous signals and ligands, along with their receptors, regulate deubiquitinase activity by protein-protein interaction. Collectively, our results provide a framework of cryptochromes and deubiquitinases to detect and interpret light signals to control plant growth at the most appropriate time.

Robust CRISPR/Cas9-Mediated Tissue-Specific Mutagenesis Reveals Gene Redundancy and Perdurance in Drosophila.

Tissue-specific loss-of-function (LOF) analysis is essential for characterizing gene function. Here, we present a simple, yet highly efficient, clustered regularly interspaced short palindromic repeats (CRISPR)-mediated tissue-restricted mutagenesis (CRISPR-TRiM) method for ablating gene function in Drosophila This binary system consists of a tissue-specific Cas9 and a ubiquitously expressed multi-guide RNA (gRNA) transgene. We describe convenient toolkits for making enhancer-driven Cas9 lines and multi-gRNAs that are optimized for mutagenizing somatic cells. We demonstrate that insertions or deletions in coding sequences more reliably cause somatic mutations than DNA excisions induced by two gRNAs. We further show that enhancer-driven Cas9 is less cytotoxic yet results in more complete LOF than Gal4-driven Cas9 in larval sensory neurons. Finally, CRISPR-TRiM efficiently unmasks redundant soluble N-ethylmaleimide-sensitive factor attachment protein receptor gene functions in neurons and epidermal cells. Importantly, Cas9 transgenes expressed at different times in the neuronal lineage reveal the extent to which gene products persist in cells after tissue-specific gene knockout. These CRISPR tools can be applied to analyze tissue-specific gene function in many biological processes.

Histone H1 defect in escort cells triggers germline tumor in Drosophila ovary.

Drosophila ovary is recognized as one of the best model systems to study stem cell biology in vivo. We had previously identified an autonomous role of the histone H1 in germline stem cell (GSC) maintenance. Here, we found that histone H1 depletion in escort cells (ECs) resulted in an increase of spectrosome-containing cells (SCCs), an ovary tumor-like phenotype. Further analysis showed that the Dpp pathway is excessively activated in these SCC cells, while the expression of bam is attenuated. In the H1-depleted ECs, both transposon activity and DNA damage had increased dramatically, followed by EC apoptosis, which is consistent with the role of H1 in other somatic cells. Surprisingly, H1-depleted ECs acquired cap cell characteristics including dpp expression, and the resulting abnormal Dpp level inhibits SCC further differentiation. Most interestingly, double knockdown of H1 and dpp in ECs can reduce the number of SCCs to the normal level, indicating that the additional Dpp secreted by ECs contributes to the germline tumor. Taken together, our findings indicate that histone H1 is an important epigenetic factor in controlling EC characteristics and a key suppressor of germline tumor.