Natural variation in the Arabidopsis thaliana circadian clock as a determinant of flowering time: a quantitative genetics and genomics study

The circadian clock is an endogenous mechanism present in plants that enables anticipation of upcoming environmental changes. In this way, the clock thus facilitates the correct timing of physiological and developmental events. An internal clock synchronized with the external environment ensures that plants flower under favorable environmental conditions, and thus, provides fitness advantages under natural habitats. In Arabidopsis, the molecular framework of the circadian-clock machinery is constituted by a network of transcription/translation-based feedback loops. The main loop that comprises CCA1/LHY and TOC1, is connected with the morning loop comprising PPR7/PRR9 through CCA1/LHY. TOC1 establishes a link of this main loop with an as of yet unknown component (Y) in the evening loop. The flowering-time component GI fulfills some of the requirement of ‘Y’. However, functional studies do not fully support the GI to TOC1 link. ELF3 is another clock component whose function is not fully understood. In the past, efforts for functional placement of ELF3 were mainly hampered by the arrhythmic nature of available elf3 alleles. In chapter 3 of this thesis, I described the characterization of a natural allele of ELF3 (ELF3-Sha) that displays conditional rhythmicity and provides evidences of the ELF3 mode-of-action in the clock. From this, in chapter 4, using quantitative genetics and genomics studies in Arabidopsis, I identified the possible positions of the new allelic variants that connect the speed of circadian oscillation to flowering time. Adaptation of Arabidopsis accessions to varying environmental conditions of light and temperature at different geographical areas has provided a force for the selection of allelic variants within the circadian clock. Previously, the location of such allelic variants affecting speed of the clock was identified in a Bay x Sha recombinant inbred line (RIL) population. This population was modified with a CCR2::LUC promoter-reporter system, which provides robust estimates of the clock activity, and hence, the speed of the clock can be accurately measured. In such experiments, a major-effect periodicity QTL was detected, where the presence of Sha allele at the locus caused a short-period phenotype. I report in chapter 3 the positional isolation of this QTL and revealed that ELF3 was the underlying gene. ELF3 has been reported to be a nuclear-localized protein required for both the generation of circadian rhythms and light input to the circadian clock. I found that the casual polymorphism in the ELF3-Sha allele was an encoded replacement in highly conserved amino acid that caused the short-period phenotype. Interestingly, this phenotype was light dependant. In constant darkness, a different phenotype was found, and this was a reduction in clock function with a movement towards arrhythmic oscillations. The cellular basis of the ELF3-Sha protein defect correlated with an increase in cytosolic distribution. Taken together, my data suggests that the light and circadian action of ELF3 in the clock is dependent on nuclear recruitment to initiate repressive action in the clock network. Characterization of ELF3-Sha established the notion that natural variation in Arabidopsis accessions provides an extraordinary resource to study complex physiological mechanisms. Therefore, I decided to exploit this natural resource to investigate the relationship between the circadian clock and flowering time, which is reported in chapter 4. For that, I selected three Arabidopsis accessions from geographically diverged areas: two from northern latitudes and one from the equator. Whole genome re-sequencing of these accessions revealed that, on a genome-wide scale, the equatorial accession is highly diverged compare to the northern accessions. This encouraged me to utilize these accessions in quantitative-genetic studies. Pair-wise crosses were made to generate RIL sets that were screened for circadian periodicity and flowering-time traits. Construction of genetic maps followed by QTL mapping revealed several loci controlling the speed of clock and flowering time. Many of these QTLs were localized with the known genes controlling both of these traits. Novel loci were also detected. Validation of a major QTL in a near isogenic line, followed by sequence comparison of the candidate gene in parental accessions, revealed that a single encoded amino-acid change in GI potentially causes the periodicity acceleration. Further, identification of flowering-time loci overlapping with the position of known clock genes provided the mechanistic explanation of the relationship of clock with flowering time. Taken together, my research strongly advocates the importance of the natural-variation studies in understanding intricate physiological pathways and their interactions with each other

Intra-radicular reinforcement using a modified fiber post customization technique: a case report

Trauma sustained by developing anterior teeth can lead to an arrested root development and loss of structure which can complicate the endodontic and restorative management. The preservation of the anterior tooth has a definite esthetic and biologic advantage especially during the developmental years of adolescence. However, restorative treatment of such cases is met with serious biomechanical and adhesive challenges in the form of thin dentinal walls, a high configuration factor etc. This case report describes a fiber post customization technique for the intraradicular reinforcement of a maxillary central incisor in a 14-year-old patient

Molecular mechanism of photoperiod sensing

ELF3 and GI are two important components of the Arabidopsis circadian clock. They are not only essential for the oscillator function but are also pivotal in mediating light inputs to the oscillator. Lack of either results in a defective oscillator causing severely compromised output pathways, such as photoperiodic flowering and hypocotyl elongation. Although single loss of function mutants of ELF3 and GI have been well-studied, their genetic interaction remains unclear. We generated an elf3 gi double mutant to study their genetic relationship in clock-controlled growth and phase transition phenotypes. We found that ELF3 and GI repress growth during the night and the day, respectively. We also provide evidence that ELF3, for which so far only a growth inhibitory role has been reported, can also act as a growth promoter under certain conditions. Finally, circadian clock assays revealed that ELF3 and GI are essential Zeitnehmers that enable the oscillator to synchronize the endogenous cellular mechanisms to external environmental signals. In their absence, the circadian oscillator fails to synchronize to the light dark cycles even under diurnal conditions. Consequently, clock-mediated photoperiod-responsive growth and development is completely lost in plants lacking both genes, suggesting that ELF3 and GI together convey photoperiod sensing to the central oscillator. Since ELF3 and GI are conserved across flowering plants and represent important breeding and domestication targets, our data highlight the possibility of developing photoperiod-insensitive crops by manipulating the combination of these two key genes

Photoperiod sensing of the circadian clock is controlled by EARLY FLOWERING 3 and GIGANTEA

ELF3 and GI are two important components of the Arabidopsis circadian clock. They are not on-ly essential for the oscillator function but are also pivotal in mediating light inputs to the oscillator. Lack of either results in a defective oscillator causing severely compromised output pathways, such as photoperiodic flowering and hypocotyl elongation. Although single loss of function mu-tants of ELF3 and GI have been well-studied, their genetic interaction remains unclear. We gen-erated an elf3 gi double mutant to study their genetic relationship in clock-controlled growth and phase transition phenotypes. We found that ELF3 and GI repress growth differentially during the night and the day, respectively. Circadian clock assays revealed that ELF3 and GI are essential Zeitnehmers that enable the oscillator to synchronize the endogenous cellular mechanisms to external environmental signals. In their absence, the circadian oscillator fails to synchronize to the light-dark cycles even under diurnal conditions. Consequently, clock-mediated photoperiod-responsive growth and development are completely lost in plants lacking both genes, suggesting that ELF3 and GI together convey photoperiod sensing to the central oscillator. Since ELF3 and GI are conserved across flowering plants and represent important breeding and domestication targets, our data highlight the possibility of developing photoperiod-insensitive crops by adjusting the allelic combination of these two key genes

Molecular Modeling Guided Drug Designing for the Therapeutic Treatment of Rheumatoid Arthritis

Rheumatoid arthritis (RA) is a systemic inflammatory disorder that can cause destructive joint disease, significant disability, and increased mortality. RA is the most frequent of all chronic inflammatory joint diseases, and its prevalence frequency in Pakistan is 1.6 per thousand people. Different cytokines and receptors were involved in the triggering of RA, including interleukin-6 (ILR-6), major histocompatibility complex (MHC) antigen human leukocyte (HLA-DR) receptor, and CD20. Several studies illustrated RA as an inherent immune response and triggered due to the "shared epitope."Therefore, the involvement of all these receptors (IL-6, HLA-DR, and CD20) leads to the neurological, ocular, respiratory, cardiac, skin, and hematological manifestations that have been considered a potential therapeutic target for drug design. Various herbal, natural, and synthetic source inhibitors of interleukin-6 (IL-6), human leukocyte (HLA-DR), and CD20 were studied and reported previously. Reported inhibitors are compared to elucidate the best inhibitor for clinical trials, leading to the orally active drug. In this study, a computer-aided drug designing approach disclosed the potential inhibitors for all receptors based on their distinct binding affinity. Moreover, drug suitability was carried out using Lipinski's rule by considering the adsorption, distribution, metabolism, and excretion (ADME) of ligands. Results elucidated "calycosin 7-O-glucoside"and "angeliferulate"as putative ligands for IL-6 and HLA-DR, respectively. However, the pharmacokinetic properties (ADMET) revealed angeliferulate as an effete ligand for the biological system compared to calycosin 7-O-glucoside. Based on docking, drug toxicity profiling or pharmacokinetics, and MD simulation stability, this study highlights orally active therapeutic inhibitors to inhibit the activity of pivotal receptors (IL6, HLA-DR, and CD20) of RA in humans. After clinical trials, the resultant inhibitors could be potential therapeutic agents in the drug development against RA

Recent developments in textile based polymeric smart sensor for human health monitoring: A review

In the modern age, the most important and prevailing issue is the monitoring of human health. To address this, several devices have been developed and a need new materials investigated. The idea of textile-based smart sensors is emerging rapidly. In this regard, ICPs and ECPs have attracted the attention of researchers due to their mechanical adaptability to suit the characteristics of textile fabric. The lighter weight, stretchability and wearability, etc. are considered an advantage while selecting the material for developing sensors not only in health monitoring but also in biomedical, sports, and military fields. The idea behind wearable sensing devices is to enable easy integration of the sensor device into daily life routines. Such wearable sensors also have the potential for real time and online monitoring of human health and integrate with smart monitoring devices. The purpose of this review is to discuss the recent developments in smart monitoring sensors.Open Access funding for this article is provided by the Qatar National Library, Al Luqta Street, Al-Rayyan P.O Box 5825 Doha, Qatar” The authors acknowledge the funding received for this work from Higher Education Commission (HEC) Pakistan under the Technology Development Fund (TDF) for grant number TDF-03-103

How Well Prepared Are Dental Students and New Graduates in Pakistan-A Cross-Sectional National Study.

The transition of an undergraduate dental student to an actual practicing dentist is a crucial phase and ensuring the preparedness of graduates for the complexity and demands of contemporary dental practice is a challenging task. This study aimed to evaluate the self-perceived preparedness of undergraduate dental students and house officers in the dental colleges of Pakistan. A cross-sectional national study was planned to collect information from dental students and new graduates in Pakistan. The pre-validated Dental Undergraduates Preparedness Assessment Scale (DU-PAS) was used. A purposive sampling technique was utilized to recruit house officers and undergraduate dental students from 27 dental schools in Pakistan. The data analysis was carried out using the R statistical environment for Windows (R Core Team, 2015). A total of 862 responses with 642 females and 219 males were analyzed in the study. Overall, the clinical skills score was 30.56 ± 9.08 and the score for soft skills was 30.54 ± 10.6. The mean age of the participants was 23.42 ± 1.28. Deficiencies were reported in various soft skills and clinical skills attributes. The results highlighted the strengths and weaknesses of dental students and new graduates in Pakistani dental institutions. The findings may be used to further develop and strengthen the teaching and training of dental students in Pakistan.The researchers would like to thank the Deanship of Scientific Research, Qassim University for funding the publication of this project

Assessment of the prevalence of radix entomolaris and distolingual canal in mandibular first molars in 15 countries: a multinational cross-sectional study with meta-analysis

Aim: The aim of this study was two-folded: i) to assess the prevalence of Distolingual Canal (DLC) and Radix Entomolaris (RE) in Mandibular First Molars (M1Ms), using Cone Beam Computed Tomography (CBCT) images and ii) to assess the impact of sociodemographic factors on the prevalence of these conditions worldwide. Methods: CBCT images were scanned retrospectively and the ones including bilateral M1Ms were included in the study. The evaluation was performed by 1 researcher in each country, trained with CBCT technology. A written and video instruction program explaining the protocol to be followed step-by-step was provided to all observers to calibrate them. The CBCT imaging screening procedure consisted of evaluating axial sections from coronal to apical. The presence of DLC and RE in M1Ms (yes/no) was identified and recorded. Results: Six thousand three hundred four CBCTs, representing 12,608 M1Ms, were evaluated. A significant difference was found between countries regarding the prevalence of both RE and DLC (P .05). Conclusion: The overall prevalence of RE and DLC in M1Ms was 3% and 22%. Additionally, both RE and DLC showed substantial bilaterally. These variations should be considered by endodontic clinicians during endodontic procedures in order to avoid potential complications

Effects of antiplatelet therapy on stroke risk by brain imaging features of intracerebral haemorrhage and cerebral small vessel diseases: subgroup analyses of the RESTART randomised, open-label trial

Background Findings from the RESTART trial suggest that starting antiplatelet therapy might reduce the risk of recurrent symptomatic intracerebral haemorrhage compared with avoiding antiplatelet therapy. Brain imaging features of intracerebral haemorrhage and cerebral small vessel diseases (such as cerebral microbleeds) are associated with greater risks of recurrent intracerebral haemorrhage. We did subgroup analyses of the RESTART trial to explore whether these brain imaging features modify the effects of antiplatelet therapy