Exploration of unique starch physicochemical properties of novel buckwheat lines created by crossing Golden buckwheat and Tatary buckwheat.

Buckwheat is considered as a healthy cereal food, and it is essential to cultivate new buckwheat lines with good starch physicochemical properties for both consumers and food producers. Six novel buckwheat (Duoku, Dk) were generated by crossing of Golden buckwheat and Tatary buckwheat, and their kernel appearance properties and starch physicochemical properties were analyzed together with one domestic line (Cimiqiao) and one wild line (Yeku). The results showed that Dk samples had better appearance properties than two control samples. The Dk samples showed lower amylose content, similar amylopectin molecular structure and chain length distributions, and larger starch granules compared with Cimiqiao. The digestion results showed that two Dk samples: Dk6 & Dk9 had high resistant starch content; while the other two Dk samples: Dk37 & Dk38 had a steady glucose releasing rate. The Dk samples also showed high gelatinization temperature, indicating they were good raw materials for producing glass noodle. This study proved that Dk buckwheat had unique starch physicochemical properties, and could be used as new food materials in the future.

UHRF1/DNMT1-MZF1 axis-modulated intragenic site-specific CpGI methylation confers divergent expression and opposing functions of PRSS3 isoforms in lung cancer.

As confusion mounts over RNA isoforms involved in phenotypic plasticity, aberrant CpG methylation-mediated disruption of alternative splicing is increasingly recognized as a driver of intratumor heterogeneity (ITH). Protease serine 3 (PRSS3), possessing four splice variants (PRSS3-SVs; PRSS3-V1-V4), is an indispensable trypsin that shows paradoxical effects on cancer development. Here, we found that PRSS3 transcripts and their isoforms were divergently expressed in lung cancer, exhibiting opposing functions and clinical outcomes, namely, oncogenic PRSS3-V1 and PRSS3-V2 versus tumor-suppressive PRSS3-V3, by targeting different downstream genes. We identified an intragenic CpG island (iCpGI) in PRSS3. Hypermethylation of iCpGI was mediated by UHRF1/DNMT1 complex interference with the binding of myeloid zinc finger 1 (MZF1) to regulate PRSS3 transcription. The garlic-derived compound diallyl trisulfide cooperated with 5-aza-2'-deoxycytidine to exert antitumor effects in lung adenocarcinoma cells through site-specific iCpGI demethylation specifically allowing MZF1 to upregulate PRSS3-V3 expression. Epigenetic silencing of PRSS3-V3 via iCpGI methylation (iCpGIm) in BALF and tumor tissues was associated with early clinical progression in patients with lung cancer but not in those with squamous cell carcinoma or inflammatory disease. Thus, UHRF1/DNMT1-MZF1 axis-modulated site-specific iCpGIm regulates divergent expression of PRSS3-SVs, conferring nongenetic functional ITH, with implications for early detection of lung cancer and targeted therapies.

Development and validation of a prognostic predictive model of pulmonary spindle cell carcinoma from the surveillance, epidemiology and end results database.

Background: Pulmonary spindle cell carcinoma (PSCC) is a rare type of non-small cell lung cancer (NSCLC). The prognostic influent factors and therapeutic methods of PSCC are unclear, for there are only some case reports or small samples' analysis. This study aims to find prognosis related factors of PSCC, develop and validate a nomogram to predict their survival probability. Methods: The Surveillance, Epidemiology, and End Results (SEER) 18 Registries database (2000-2018) was searched to study PSCC. According to diagnosed time, data was divided into primary cohort (2000-2015) and validation cohort (2016-2018), both followed until December 31 2018. Chosen by Least Absolute Shrinkage and Selection Operator (LASSO) regression, age, sex, stage, surgery, chemotherapy, N, size and history of malignancy were taken out as predictive variables. The primary cohort was used to develop a nomogram to predict 1-, 3- and 5-year overall survival (OS) probability, and be validated by the validation cohort using concordance index (C-index) and calibration curves. Both cohorts were used to conduct a Cox regression to find the influential factors on OS of PSCC. Results: The nomogram shows a good concordance and discrimination on the prediction of OS, both internal (n=457 and C-index is 0.79) and external validation (n=100 and C-index is 0.76). The median survival time of PSCC is 4 months, with 20.1% OS possibility in 5 years. Multivariate analysis identified patients of older age [hazard ratio (HR), 1.02; 95% confidence interval (CI): 1.01-1.04], larger size of neoplasm (HR, 1.01; 95% CI: 1.01-1.01), M1 (HR, 2.96; 95% CI: 2.17-4.04), N2 (HR, 2.55; 95% CI: 1.81-3.59) or N3 (HR, 2.99; 95% CI: 1.58-5.66), regional stages (HR, 2.11; 95% CI: 1.29-3.44) and distant stages (HR, 6.17; 95% CI: 3.83-9.94) had a lower OS possibility, while surgery (HR, 0.39; 95% CI: 0.28-0.53) and history of malignancy (HR, 0.68; 95% CI: 0.48-0.98) was protective factors for PSCC. PSCC survived longer with surgery performed instead of chemotherapy or radiotherapy. Conclusions: Patients of PSCC have a poor prognosis, and using the nomogram developed by this study can predict their 1-, 3- and 5-year OS probability. Surgery is a better choice for PSCC and more studies are necessary to find potential treatment like targeted therapy, programmed death-1 (PD-1) and programmed death ligand 1 (PD-L1).

Long non-coding RNA BZRAP1-AS1 functions in malignancy and prognosis for non-small-cell lung cancer.

Purpose: The function of BZRAP1-AS1 is unknown in lung cancer. We evaluated the clinicopathologic significance of BZRAP1-AS1, and its role in non-small-cell lung cancer (NSCLC) progression. Patient and methods: Sixty-three NSCLC patients from Beijing Chest Hospital were included. The expression of BZRAP1-AS1 was detected by real-time quantitative polymerase chain reaction (RT-qPCR) in tumor tissues and adjacent normal tissues. Then, the clinicopathological significance and prognostic value of BZRAP1-AS1 were analyzed by using our cohort and TCGA cohort. Finally, the effect of BZRAP1-AS1 on proliferation and motility of NSCLC cell lines were evaluated by cell growth assay, colony formation assay, xenograft tumorigenesis experiment in nude mice and transwell assays respectively. Results: Compared with adjacent normal tissues, BZRAP1-AS1 showed lower expression in NSCLC tumor tissues. As for the relationship between BZRAP1-AS1 and clinical characteristics, our results were consistent with those of TCGA data. BZRAP1-AS1 was lower in T1 than T2-T4 patients, N1-N3 than N0 patients. Low level BZRAP1-AS1 was related to shorter overall survival time (OS) in lung adenocarcinoma (LUAD), and poor first progression time (FP) in LUAD and lung squamous cell carcinoma (LUSC) patients. BZRAP1-AS1 was significantly associated with the prognosis of NSCLC patients. Overexpression of BZRAP1-AS1 inhibited proliferation and migration of H1299 and HCC827 cells. Conclusion: BZRAP1-AS1 expression decreases in tumor tissues with the increase of malignancy grades in NSCLC. BZRAP1-AS1 plays an anticancer role by inhibiting cell proliferation, invasion, and metastasis, and has a potential prognostic value in NSCLC. BZRAP1-AS1 may serve as a diagnostic marker and therapeutic target for NSCLC.

Re-evaluating the need for mediastinal lymph node dissection and exploring lncRNAs as biomarkers of N2 metastasis in T1 lung adenocarcinoma.

Background: Although a well-acknowledged component of curative surgery for lung cancer, investigators have recently questioned the need for mediastinal lymph node dissection (MLND) in early-stage lung cancer cases. As such, the accurate prediction of N2 stage prior to surgery has become increasingly critical. But diagnostic biomarkers predicting N2 metastases are deficient, which are urgently needed. Methods: We extracted the data of non-small cell lung cancer (NSCLC) patients whose clinical information and follow-up data are complete and without preoperative induction therapy from the Surveillance, Epidemiology, and End Results (SEER) database. The SEER program registries routinely collect demographic and clinic data on patients. And the prognostic differences were analyzed according to the presence or absence of MLND in their lung resection using the R package. Subsequently, the correlations between pN2 metastasis and clinical characteristics were analyzed. In parallel, the long non-coding RNAs (lncRNAs) associated with pN2 status were screened in The Cancer Genome Atlas (TCGA) database by expression difference analysis between pN0-N1 and pN2 patients using limma. Their diagnostic efficiency for detecting N2 metastases was evaluated using receiver operating characteristic (ROC) curves, and a combined diagnostic model was constructed using logistic regression and ROC curve analyses in lung adenocarcinoma (LUAD). Results: There were 16,772 patients in MLND group, and 2,699 cases in no-MLND group. The clinical data from SEER showed that the incidence of N2 metastasis was low in pT1 NSCLC (1,023/16,772, 6.10%), but the prognosis of no-MLND patients was poorer than those who underwent MLND (P<0.001, HR =1.605). Pathological N2 metastasis was correlated with age, histologic type, and tumor size. On the other hand, five lncRNAs (LINC00892, AC099522.2, LINC01481, SCAMP1-AS1, and AC004812.2) were screened and confirmed as potential diagnostic biomarkers for detecting N2 metastasis in pT1 LUAD. The AUC of the combined indicators was 0.857. Conclusions: MLND may be oncologically necessary for selected T1 NSCLC patients based on the metastasis incidence and prognosis. A diagnostic model combining LINC00892, AC099522.2, LINC01481, SCAMP1-AS1, and AC004812.2 expression levels may have the potential to be a diagnostic biomarker for detecting N2 metastasis in pT1 LUAD. This study suggests that MLND might be omitted in patients with lower expression level of this diagnostic model.

[Analysis of the Efficacy of Immunotherapy in Elderly Patients with Lung Cancer].

BACKGROUND: Immunotherapy represented by immune checkpoint inhibitors (ICIs) has become the standard treatment for patients with non-oncogenic advanced non-small cell lung cancer (NSCLC). While lung cancer is most prevalent in elderly patients, these patients are rarely included in pivotal clinical trial studies. We aimed to describe the efficacy and safety of immunotherapy for elderly patients in the "real-world". METHODS: The data of older NSCLC patients and younger patients who received immunotherapy between July 2018 to October 2021 were retrospectively analyzed and the objective response rate (ORR) and progression-free survival (PFS) in different age groups (less than 60 years old was defined as the young group, 60 years-74 years old was the young old group, 75 years old and above was the old old group) were compared. And the impact of different clinical characteristics on treatment response and prognosis were analyzed in each age subgroup. RESULTS: A total of 21 young patients, 70 young old patients and 15 old old patients were included in this study, with ORR of 33.3%, 52.8% and 53.3%, respectively, without statistically significant difference (P=0.284). The median PFS was 9.1 mon, 7.6 mon and 10.9 mon, respectively, without statistically significant difference (P=0.654). Further analysis of the predictors of immunotherapy in each subgroup revealed that patients in the young old group and young group who received immunotherapy in the first line had a longer PFS. The difference of the incidence of adverse events was not statistically significant among the three groups (P>0.05). CONCLUSIONS: The efficacy and safety of immunotherapy in elderly patients were similar to those in younger patients, and PFS was superior in the first-line immunotherapy. Further prospective studies are still needed to explore predictors of immunotherapy in elderly NSCLC patients.

Early ligation of the pulmonary vein can reduce the dissemination of shed tumor cells during thoracoscopic lobectomy.

OBJECTIVE: The sequence of vessel ligation in lobectomy can significantly affect the hematogenous spread of circulating tumor cells (CTCs). Vein-first ligation substantially reduces CTC dissemination and achieves favorable survival compared with artery-first ligation. In this study, we further explored whether the timing of pulmonary vein (PV) ligation determined according to the early and late PV ligation technique is associated with CTC dissemination. METHODS: A total of 44 patients who underwent uniform 2-port video-assisted thoracoscopic surgery lobectomy were enrolled; the subjects were divided into the early ligation group (n = 18) and late ligation group (n = 26) according to whether PV ligation was prioritized during surgery. PV blood was obtained before PV ligation and after lobe resection. CTCs were detected using telomerase reverse transcriptase-based CTC detection and validated using FlowSight and fluorescence in situ hybridization. RESULTS: The median postoperative PV CTC (Post-PVCTC) count was 9 (interquartile range [IQR], 6-18), which was higher than the median preoperative PV CTC (Pre-PVCTC) count of 1 (IQR, 0-3; P < .001). Clinicopathologic correlation analysis showed that the Pre-PVCTC count correlated positively with TNM stage (P = .002) and lymph node metastasis (P = .002) and that the Post-PVCTC count correlated positively with tumor density (P = .043) and vessel/lymphatic invasion (P < .030). Interestingly, although no statistical difference in the median Pre-PVCTC count was observed, the median Post-PVCTC count in the early ligation group was 16 (IQR, 9.5-36.75), whereas that in the late ligation group it was 8 (IQR, 4.75-12.25), showing a significant difference (P = .004). CONCLUSIONS: We provide the first evidence to show that early PV ligation can prevent PVCTCs from spreading into the circulation, offering an innovative surgical concept for the principle sequence of pulmonary vessel management.

H2S aids osmotic stress resistance by S-sulfhydration of melatonin production-related enzymes in Arabidopsis thaliana.

KEY MESSAGE: Hydrogen sulfide closed Arabidopsis thaliana stomata by increasing the transcription of melatonin-producing enzymes and the post-translational modification levels to combat osmotic stress. Hydrogen sulfide (H2S) and melatonin (MEL) reportedly have similar functions in many aspects of plant growth, development and stress response. They regulate stomatal movement and enhance drought resistance. However, their physiological relationship is not well understood. Here, their crosstalk involved in osmotic stress resistance in Arabidopsis thaliana was studied. Exogenous H2S and MEL closed stomata under normal or osmotic stress conditions and increased the relative water contents of plants under osmotic stress conditions. At the same time, exogenous H2S and MEL responded to osmotic stress by increasing the content of proline and soluble sugar, and reducing malondialdehyde (MDA) content and relative conductivity. Using mutants in the MEL-associated production of serotonin N-acetyltransferase (snat), caffeic acid O-methyltransferase (comt1) and N-acetylserotonin methyltransferase (asmt), we determined that H2S was partially dependent on MEL to close stomata. Additionally, the overexpression of ASMT promoted stomatal closure. Exogenous H2S increased the transcription levels of SNAT, ASMT and COMT1. Furthermore, exogenous H2S treatments increased the endogenous MEL content significantly. At the post-translational level, H2S sulfhydrated the SNAT and ASMT, but not COMT1, enzymes associated with MEL production. Thus, H2S appeared to promote stomatal closure in response to osmotic stress by increasing the transcription levels of MEL synthesis-related genes and the sulfhydryl modification of the encoded enzymes. These results increased our understanding of H2S and MEL functions and interactions under osmotic stress conditions.

Subsegmentectomy versus segmentectomy resection for the treatment of operable patients with stage IA non-small cell lung cancer: A meta-analysis.

Background: There were new points of interest in performing subsegmentectomy and segmentectomy for patients with early stage non-small cell lung cancer (NSCLC). However, whether patients who underwent subsegmentectomy could obtain satisfactory clinical outcomes remains unclear. The present study aimed to compare the clinical outcomes and security of surgical procedures between subsegmentectomy and segmentectomy. Methods: A systematic review and meta-analysis was performed through five online databases to identify the included literatures which presented intact clinical outcome data among different surgical procedures. The included studies were evaluated based on precise and predefined inclusion criteria. Results: There were 4 published studies identified in this meta-analysis. A total of 325 patients who underwent subsegmentectomy and 904 patients who underwent segmentectomy were involved in this analysis. The duration of drainage [MD -0.19; 95%CI (-0.36, -0.02), p = 0.03] and postoperative hospital stay [MD -0.30; 95%CI (-0.58, -0.02), p = 0.009] of subsegmentectomy were significantly less than that of segmentectomy. There was no statistically significant difference among recurrence rate [OR 0.85; 95%CI (0.21, 3.42), p = 0.82], operation time, blood loss, incidence of complications [OR 0.83; 95%CI (0.58, 1.20), p = 0.33] between subsegmentectomy and segmentectomy in patients with stage IA NSCLC. Conclusion: The meta-analysis was firstly performed to compare perioperative outcomes among surgical procedures. The perioperative outcomes were comparable between subsegmentectomy and segmentectomy. Subsegmentectomy might be an alternative treatment for the deep tumor with size less than 1.5 cm and mainly composed of Ground Glass Opacity (GGO).

[Preliminary Efficacy Evaluation of Neoadjuvant Immunotherapy Combined with 
Chemotherapy in Resectable Non-small Cell Lung Cancer].

BACKGROUND: Preliminary researches conformed that neoadjuvant immunotherapy combined with chemotherapy had a significant short-term effect in resectable non-small cell lung cancer (NSCLC), but there were few clinical trials about neoadjuvant chemoimmunotherapy in China. We aimed to assess retrospectively the antitumour activity and safety of neoadjuvant chemoimmunotherapy for resectable stage Ib-IIIb NSCLC. METHODS: Twenty patients who had been diagnosed as stage Ib-IIIb NSCLC and received chemoimmunotherapy as neoadjuvant treatment between November 2019 and December 2020, in Beijing Chest Hospital, Capital Medical University were recruited. These patients received neoadjuvant treatment for 21 days as a cycle and antitumour activity and safety were evaluated every two cycles. RESULTS: Of 20 patients received neoadjuvant chemoimmunotherapy, 17 patients underwent surgical resection. 16 patients had R0 resection (no residual tumor resection) and 1 patient had R1 resection (microscopic residual tumor resection). Radiographic objective response rate (ORR) was 85.0% (4 complete response, 13 partial response). 5.0% (1/20) of patients had stable disease, and 10.0% (2/20) of patients had progression disease. The major pathologic response (MPR) was 47.1% (8/17), and complete pathologic response (CPR) was 29.4% (5/17). 1 case developed grade IV immune-related pneumonia (IRP) and 9 (45.0%) cases had grade III hematologic toxicity. CONCLUSIONS: Immunotherapy combined with chemotherapy as neoadjuvant therapy has a better efficiency and tolerable adverse effects for patients with resectable NSCLC in stage Ib-IIIb.

Clinical Significance of miR-183-3p and miR-182-5p in NSCLC and Their Correlation.

PURPOSE: Accumulating evidence has indicated that dysregulated microRNAs (miRNAs) are involved in cancer progression. In this study, we evaluated the clinicopathologic significance of miR-183-3p and miR-182-5p, and the role of miR-183-3p in non-small-cell lung cancer (NSCLC) progression. PATIENTS AND METHODS: Seventy-six NSCLC patients from Beijing Chest Hospital were included. The expression of miR-183-3p and miR-182-5p was evaluated by real-time quantitative polymerase chain reaction (RT-qPCR). Then, cell growth curve assays and colony formation assays were performed. Bioinformatics analysis of TCGA database was performed to explore the clinicopathological significance and prognostic value. RESULTS: miR-183-3p and miR-182-5p were significantly increased in NSCLC tumor tissues (both P < 0.0001) and were positively correlated (r = 0.8519, P < 0.0001). miR-183-3p (P = 0.0444) and miR-182-5p (P = 0.0132) were correlated with tumor size. In addition, miR-183-3p (P = 0.0135) and miR-182-5p (P = 0.0009) were upregulated in normal lung tissues from smokers. In vitro, miR-183-3p was correlated with cell proliferation. In addition, bioinformatics analysis indicated that miR-183-3p was correlated with poor prognosis (P = 0.0466) and tumor size (P = 0.0017). In addition, miR-183-3p was higher in lung squamous carcinoma (LUSC) tissue (P < 0.0001) than in lung adenocarcinoma (LUAD) tissue, and miR-183-3p was higher in the tumor tissue of smokers (P = 0.0053) than in that of nonsmokers. CONCLUSION: Upregulation of miR-183-3p and miR-182-5p may play an oncogenic role in NSCLC. miR-183-3p could be used as a potential prognostic biomarker and therapeutic target to manage lung cancer.

Hydrogen sulfide promotes flowering in heading Chinese cabbage by S-sulfhydration of BraFLCs.

Heading Chinese cabbage (Brassica rapa L. syn. B. campestris L. ssp. chinensis Makino var. pekinensis (Rupr.) J. Cao et Sh. Cao) is a cruciferous Brassica vegetable that has a triplicate genome, owing to an ancient genome duplication event. It is unclear whether the duplicated homologs have conserved or diversified functions. Hydrogen sulfide (H2S) is a plant gasotransmitter that plays important physiological roles in growth, development, and responses to environmental stresses. The modification of cysteines through S-sulfhydration is an important mechanism of H2S, which regulates protein functions. H2S promotes flowering in Arabidopsis and heading Chinese cabbage. Here we investigated the molecular mechanisms of H2S used to promote flowering in the latter. Four, five, and four BraFLC, BraSOC I, and BraFT homologs were identified in heading Chinese cabbage. Different BraFLC proteins were bound to different CArG boxes in the promoter regions of the BraSOC I and BraFT homologs, producing different binding patterns. Thus, there may be functionally diverse BraFLC homologs in heading Chinese cabbage. Exogenous H2S at 100 µmol L-1 significantly promoted flowering by compensating for insufficient vernalization. BraFLC 1 and BraFLC 3 underwent S-sulfhydration by H2S, after which their abilities to bind most BraSOC I or BraFT promoter probes weakened or even disappeared. These changes in binding ability were consistent with the expression pattern of the BraFT and BraSOC I homologs in seedlings treated with H2S. These results indicated that H2S signaling regulates flowering time. In summary, H2S signaling promoted plant flowering by weakening or eliminating the binding abilities of BraFLCs to downstream promoters through S-sulfhydration.

Effects of lithium resource exploitation on surface water at Jiajika mine, China.

Heavy metal and metalloid (HMM) contamination of the water environment caused by mining activities is a great challenge to the global mining industry. HMMs released by various mines could easily enter the surrounding environment and pose serious threats to human health. Although the HMM pollution of surface water in various mines has been widely researched, relevant studies on the effects of mining activities on the surface water of hard-rock-type Li mines are scarce. Herein, a total of 81 water samples were collected from Jiajika mine for the first time, the largest hard-rock-type Li mine in Asia. The physical parameters and concentrations of HMMs and major ions of the samples were analyzed to evaluate the water quality and HMM level of surface water. Results showed that (1) most of the parameters analyzed adhered to the strictest guidelines of Chinese surface waters and the drinking water guidelines of WHO, except Mn, Pb, and As of a few samples from tailings-affected areas and Li-bearing areas; (2) mineral tailings obviously increased the pH and decreased the dissolved oxygen (DO) of the surrounding surface waters; (3) the highest concentrations of As (5.58 µg/L), Zn (81.8 µg/L), Ba (5.26 µg/L), and Co (0.33 µg/L) were observed around the tailings reservoir, whereas the highest concentrations of Cr (1.5 µg/L), Mn (380 µg/L), Pb (28.4 µg/L), and V (3.16 µg/L) were observed in Li-bearing areas; and (4) according to the statistical results, the concentrations of As, Cr, Ni, and V in surface water were mainly affected by mining activities, whereas those of Cu, Zn, Ba, Co, and Pb were dominantly affected by natural processes. These results provide useful information about water quality in relation to Li mining and can help the government make reasonable decisions regarding hard-rock-type Li resource exploitation activities.

Cloning and expression analysis of a new anther-specific gene CaMF4 in Capsicum annuum.

Our previous study on the genic male sterile-fertile line 114AB of Capsicum annuum indicated a diversity of differentially expressed cDNA fragments in fertile and sterile lines. In this study, a transcript-derived fragment (TDF), male fertile 4 (CaMF4) was chosen for further investigation to observe that this specific fragment accumulates in the flower buds of the fertile line. The full genomic DNA sequence of CaMF4 was 894 bp in length, containing two exons and one intron, and the complete coding sequence encoded a putative 11.53 kDa protein of 109 amino acids. The derived protein of CaMF4 shared similarity with the members of PGPS/D3 protein family. The expression of CaMF4 was detected in both the flower buds at stage 8 and open flowers of the male fertile line. In contrast to this observation, expression of CaMF4 was not detected in any organs of the male sterile line. Further analysis revealed that CaMF4 was expressed particularly in anthers of the fertile line. Our results suggest that CaMF4 is an anther-specific gene and might be indispensable for anther or pollen development in C. annuum.

Isolation and characterization of CaMF3, an anther-specific gene in Capsicum annuum L.

Previous work on gene expression analysis based on RNA sequencing identified a variety of differentially expressed cDNA fragments in the genic male sterile-fertile line 114AB of Capsicum annuum L. In this work, we examined the accumulation of one of the transcript-derived fragments (TDFs), CaMF3 (male fertile 3), in the flower buds of a fertile line. The full genomic DNA sequence of CaMF3 was 1,951 bp long and contained 6 exons and 5 introns, with the complete sequence encoding a putative 25.89 kDa protein of 234 amino acids. The predicted protein of CaMF3 shared sequence similarity with members of the isoamyl acetate-hydrolyzing esterase (IAH1) protein family. CaMF3 expression was detected only in flower buds at stages 7 and 8 and in open flowers of a male fertile line; no expression was observed in any organs of a male sterile line. Fine expression analysis revealed that CaMF3 was expressed specifically in anthers of the fertile line. These results suggest that CaMF3 is an anther-specific gene that may be essential for anther or pollen development in C. annuum.

CaMF2, an anther-specific lipid transfer protein (LTP) gene, affects pollen development in Capsicum annuum L.

Based on the gene differential expression analysis performed by cDNA-amplified fragment length polymorphism (cDNA-AFLP) in the genic male sterile-fertile line 114AB of Capsicum annuum L., a variety of differentially expressed cDNA fragments were detected in fertile or sterile lines. A transcript-derived fragment (TDF) specifically accumulated in the flower buds of fertile line was isolated, and the corresponding full-length cDNA and DNA were subsequently amplified. Bioinformatical analyses of this gene named CaMF2 showed that it encodes a lipid transfer protein with 94 amino acids. Spatial and temporal expression patterns analysis indicated that CaMF2 was an anther-specific gene and the expression of CaMF2 was detected only in flower buds at stage 3-7 of male fertile line with a peak expression at stage 4, but not detected in the roots, tender stems, fresh leaves, flower buds, open flowers, sepals, petals, anthers or pistils of male sterile line. Further, inhibition of the CaMF2 by virus-induced gene silencing (VIGS) method resulted in the low pollen germination ability and shriveled pollen grains. All these evidence showed that CaMF2 had a vital role in pollen development of C. annuum.