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1.
Genes (Basel) ; 14(2)2023 01 30.
Article in English | MEDLINE | ID: mdl-36833291

ABSTRACT

Identifying the ideal plant nature and canopy structure is of great importance for improving photosynthetic production and the potential action of plants. To address this challenge, an investigation was accomplished in 2018 and 2019 at the Institute of Cotton Research (ICR) of the Chinese Academy of Agricultural Science (CAAS), Henan Province, China. Six cotton varieties with diverse maturities and plant canopy structures were used to evaluate the light interception (LI) in cotton, the leaf area index (LAI), the biomass, and the yield throughout the two years of study. The light spatial distribution in the plant canopy was evaluated using a geographic statistical method, following the increasing quantity of radiation intercepted, which was determined using the rules of Simpson. Compared to the cotton plants with a compact structure, varieties with both a loose and tower design captured a comparatively higher amount of light (average 31.3%) and achieved a higher LAI (average 32.4%), eventually achieving a high yield (average 10.1%). Furthermore, the polynomial correlation revealed a positive relationship between the biomass accumulation in the reproductive parts and canopy-accrued light interception (LI), signifying that light interception is critical for the yield development of cotton. Furthermore, when the leaf area index (LAI) was peaked, radiation interception and biomass reached the highest during the boll-forming stage. These findings will provide guidance on the light distribution in cotton cultivars with an ideal plant structure for light capture development, providing an important foundation for researchers to better manage light and canopies.


Subject(s)
Gossypium , Photosynthesis , Biomass , Agriculture , Plant Leaves
2.
Sci Total Environ ; 821: 153407, 2022 May 15.
Article in English | MEDLINE | ID: mdl-35090924

ABSTRACT

Analyzing the carbon footprint of crop production and proposing low-carbon emission reduction production strategies can help China develop sustainable agriculture under the goal of 'carbon peak and carbon neutrality'. Cotton is an economically important crop in China, but few reports have systematically quantified the carbon footprint of China's cotton production and analyzed its spatiotemporal changes and driving factors. This study used a life cycle approach to analyze the spatiotemporal changes and identify the main components and driving factors of the carbon footprint of cotton production in China between 2004 and 2018 based on statistical data. The results showed that the carbon footprint per unit area of cotton in Northwest China, the Yellow River Basin and the Yangtze River Basin reached 6220.13 kg CO2eq·ha-1, 3528.14 kg CO2eq·ha-1 and 2958.56 kg CO2eq·ha-1, respectively. From 2004 to 2018, the CFa in the Yellow River Basin and Northwest China increased annually, with average increases of 59.87 kg CO2eq·ha-1 and 260.70 kg CO2eq·ha-1, respectively, while the CFa in the Yangtze River Basin decreased by an average of 21.53 kg CO2eq·ha-1 per year. The ridge regression and Logarithmic Mean Divisia Index (LMDI) model showed that fertilizer, irrigation electricity and agricultural film were the main influences on carbon emission growth at the micro level and that the economic factor was the key factor at the macro level. Improving the efficiency of cotton fertilization and electricity use and ensuring the high-quality development of the cotton industry are effective strategies to reduce the carbon footprint of cotton cultivation in the future. This study comprehensively uses statistical data and mathematical modeling to provide theoretical support for accounting and in-depth analysis of cotton carbon emissions. The results are valuable for policy making related to sustainable development and the low-carbon development of the Chinese cotton industry.


Subject(s)
Carbon Footprint , Fertilizers , Agriculture/methods , Carbon/analysis , China , Fertilizers/analysis , Rivers
3.
JOM (1989) ; 73(6): 1696-1704, 2021 Jun.
Article in English | MEDLINE | ID: mdl-34456537

ABSTRACT

Amelogenin is the most abundant matrix protein guiding hydroxyapatite formation in enamel, the durable bioceramic tissue that covers vertebrate teeth. Here, we sought to refine structure-function for an amelogenin domain based on in vitro data showing a 42 amino acid amelogenin-derived peptide (ADP7) mimicked formation of hydroxyapatite similar to that observed for the full-length mouse 180 amino acid protein. In mice, we used CRISPR-Cas9 to express only ADP7 by the native amelogenin promoter. Analysis revealed ADP7 messenger RNA expression in developing mouse teeth with the formation of a thin layer of enamel. In vivo, ADP7 peptide partially replaced the function of the full-length amelogenin protein and its several protein isoforms. Protein structure-function relationships identified through in vitro assays can be deployed in whole model animals using CRISPR-Cas9 to validate function of a minimal protein domain to be translated for clinical use as an enamel biomimetic.

4.
Ying Yong Sheng Tai Xue Bao ; 32(3): 1112-1118, 2021 Mar.
Article in English | MEDLINE | ID: mdl-33754579

ABSTRACT

Cotton is one of the most important crops in the world. With the increasing scarce of global water resources, irrigation water will become a major limiting factor in cotton production. Deficit irrigation is an irrigation method which consumes less water than the normal evapotranspiration of crops. It is an effective water-saving method due to improved water use efficiency without sacrificing cotton yield and fiber quality. We summarized the effects of deficit irrigation on the growth and water use efficiency of cotton. The results showed that deficit irrigation promoted the transformation from vegetative growth to reproductive growth, reduced plant height, leaf area, and total biomass of cotton, and subsequently improved the harvest index, stem diameter and water use efficiency. Finally, based on the current research and combined with cotton production reality, the application and future development of deficit irrigation were proposed, which might provide theoretical guidance for the sustainable development of cotton plantation in arid areas.


Subject(s)
Agricultural Irrigation , Water , Biomass , Crops, Agricultural , Plant Leaves
5.
PLoS One ; 14(9): e0222395, 2019.
Article in English | MEDLINE | ID: mdl-31557178

ABSTRACT

The growth and development of cotton are closely related to climatic variables such as temperature and solar radiation. Adjusting planting density is one of the most effective measures for maximizing cotton yield under certain climatic conditions. The objectives of this study were (1) to determine the optimum planting density and the corresponding leaf area index (LAI) and yield under the climatic conditions of Henan Province, China, and (2) to learn how climatic conditions influence cotton growth, yield, and yield components. A three-year (2013-2015) field experiment was conducted in Anyang, Henan Province, using cultivar SCRC28 across six planting density treatments: 15,000, 33,000, 51,000, 69,000, 87,000, and 105,000 plants ha-1. The data showed that the yield attributes, including seed cotton yield, lint yield, dry matter accumulation, and the LAI, increased as planting density increased. Consequently, the treatment of the maximum density with 105,000 plants ha-1 was the highest-yielding over three years, with the LAIs averaged across the three years being 0.37 at the bud stage, 2.36 at the flower and boll-forming stage, and 1.37 at the boll-opening stage. Furthermore, the correlation between the cotton yield attributes and meteorological conditions indicated that light interception (LI) and the diurnal temperature range were the climatic factors that most strongly influenced cotton seed yield. Moreover, the influence of the number of growing degree days (GDD) on cotton was different at different growth stages. These observations will be useful for determining best management practices for cotton production under the climatic conditions of Henan Province, China.


Subject(s)
Crop Production/methods , Gossypium , China , Climate , Crop Production/statistics & numerical data , Gossypium/growth & development
6.
Spectrochim Acta A Mol Biomol Spectrosc ; 221: 117184, 2019 Oct 05.
Article in English | MEDLINE | ID: mdl-31158773

ABSTRACT

Silver nanowire arrays as surface-enhanced Raman scattering (SERS) substrates were prepared by a solid-state ionics method under the direct current electric field (DCEF) and used to rapidly detect melamine in aqueous solutions. The arrangement density and surface roughness of the prepared silver nanowire arrays are significantly different upon a change in the impressed current intensity. The growth mechanism of silver nanowire arrays was associated with the apical growth advantage and the irregular electrode interface. When the current intensity was 4 µA and 10 µA, the fractal dimension of silver nanowire arrays was 1.66 and 1.49, the diameters of nanowires ranged from 90 to 130 nm and 90 to 170 nm, and many densely arranged and regularly arranged silver nanoparticles lie in the prepared nanowire arrays, respectively. The result shows that there were more silver nanostructures and surface roughness under 4 µA DCEF. The Raman signal intensity of melamine molecule shows that the prepared SERS substrate exhibited a high sensitivity. The proposed method allow us detect melamine with a limit of 10-15 mol/L and 10-12 mol/L, which are lower than the safety limit estimated by the US food and Drug Administration. With its facile material synthesis, simple detection procedure and low detection concentration, this silver nanowire arrays with high surface roughness indicates a strong potential detection technique in the field of food safety.


Subject(s)
Nanowires/chemistry , Silver/chemistry , Spectrum Analysis, Raman/methods , Triazines/analysis , Food Analysis/instrumentation , Food Analysis/methods , Food Contamination/analysis , Fractals , Limit of Detection , Sensitivity and Specificity , Solutions/analysis , Spectrum Analysis, Raman/instrumentation , Vibration , Water/chemistry
7.
PLoS One ; 14(5): e0217243, 2019.
Article in English | MEDLINE | ID: mdl-31107925

ABSTRACT

Different cotton (Gossypium hirsutum L.)-wheat (Triticum aestivum) planting patterns are widely applied in the Yellow River Valley of China, and crop yield mainly depends on light interception. However, little information is available on how cotton canopy light capturing and yield distribution are affected by planting patterns. Hence, field experiments were conducted in 2016 and 2017 to study the response of cotton canopy light interception, square and boll distribution, the leaf area index (LAI) and biomass accumulation to three planting patterns: a cotton monoculture (CM, planted on 15 May) system, a cotton/wheat relay intercropping (CWI, planted on 15 May) system, in which three rows of wheat rows were intercropped with one row of cotton, and a system in which cotton was directly seeded after wheat (CWD, planted on 15 June). The following results were obtained: 1) greater light capture capacity was observed for cotton plants in the CM and CWI compared with the CWD, and the light interception of the CM was 22.4% and 51.4% greater than that of the CWI and CWD, respectively, at 30 days after sowing (DAS) in 2016; 2) more bolls occurred at the first sympodial position (SP) than at other SPs for plants in the CM; 3) based on the LAI and biomass accumulation, the cotton growth rate was the greatest in CWD, followed by CM and CWI; and 4) the CM produced significantly greater yields than did the other two treatments because it yielded more bolls and greater boll weight. Information on the characteristics of cotton growth and development in response to different planting patterns would be helpful for understanding the response of cotton yields to planting patterns and would facilitate the improvement of cotton productivity.


Subject(s)
Crop Production/methods , Gossypium/growth & development , Triticum/growth & development , Biomass , Carbohydrate Metabolism , China , Crops, Agricultural/growth & development , Edible Grain/growth & development , Gossypium/metabolism , Gossypium/radiation effects , Light , Plant Leaves/growth & development , Seeds/growth & development , Triticum/metabolism , Triticum/radiation effects
8.
PLoS One ; 12(12): e0190032, 2017.
Article in English | MEDLINE | ID: mdl-29272298

ABSTRACT

In this study, transplanting cotton seedlings grown in artificial substrate is considered due to recent increased interest in cotton planting labor saving approaches. The nursery methods used for growing cotton seedlings affect root growth. However, the underlying functional responses of root growth to variations in cotton seedling transplanting methods are poorly understood. We assessed the responses of cotton (Gossypium hirsutum L.) roots to different planting methods by conducting cotton field experiments in 2012 and 2013. A one-factor random block design was used with three replications and three different cotton planting patterns (substrate seedling transplanted cotton (SSTC), soil-cube seedling transplanted cotton (ScSTC) and directly sown cotton (DSC). The distributions and variances of the root area density (RAD) and root length density (RLD) at different cotton growing stages and several yield components were determined. Overall, the following results were observed: 1) The RAD and RLD were greatest near the plants (a horizontal distance of 0 cm) but were lower at W20 and W40 cm in the absence of film mulching than at E20 and E40 cm with film mulching. 2) The roots were confined to shallow depths (20-40 cm), and the root depths of SSTC and DSC were greater than the root depths of ScSTC. 3) Strong root growth was observed in the SSTC at the cotton flowering and boll setting stages. In addition, early onset root growth occurred in the ScSTC, and vigorous root growth occurred throughout all cotton growth stages in DSC. 4) The SSTC plants had more lateral roots with higher root biomass (RB) than the ScSTC, which resulted in higher cotton yields. However, the early onset root growth in the ScSTC resulted in greater pre-frost seed cotton (PFSC) yields. These results can be used to infer how cotton roots are distributed in soils and capture nutrients.


Subject(s)
Gossypium/growth & development , Plant Roots/growth & development , Seedlings/growth & development , Biomass
9.
PLoS One ; 12(3): e0172537, 2017.
Article in English | MEDLINE | ID: mdl-28257469

ABSTRACT

As an important successional stage and main type of biological soil crusts (BSCs) in Shapotou region of China (southeastern edge of Tengger Desert), lichen soil crusts (LSCs) often suffer from many stresses, such as desiccation and excess light intensity. In this study, the chlorophyll fluorescence and CO2 exchange in the rehydrated LSCs were detected under a series of photosynthetically active radiation (PAR) gradients to study the photosynthetic acclimation of LSCs. The results showed that although desiccation leaded to the loss of photosynthetic activity in LSCs, the fluorescence parameters including Fo, Fv and Fv/Fm of LSCs could be well recovered after rehydration. After the recovery of photosynthetic activity, the effective photosynthetic efficiency ΦPSII detected by Imaging PAM had declined to nearly 0 within both the lichen thallus upper and lower layers when the PAR increased to 200 µE m-2 s-1, however the net photosynthesis detected by the CO2 gas analyzer in the LSCs still appeared when the PAR increased to 1000 µE m-2 s-1. Our results indicate that LSCs acclimating to high PAR, on the one hand is ascribed to the special structure in crust lichens, making the incident light into the lichen thallus be weakened; on the other hand the massive accumulation of photosynthetic pigments in LSCs also provides a protective barrier for the photosynthetic organisms against radiation damage. Furthermore, the excessive light energy absorbed by crust lichens is also possibly dissipated by the increasing non-photochemical quenching, therefore to some extent providing some protection for LSCs.


Subject(s)
Cyanobacteria/physiology , Lichens/physiology , Photosynthesis/physiology , Soil Microbiology , Acclimatization/physiology , China , Chlorophyll/chemistry , Desert Climate , Ecosystem , Light , Temperature
10.
PLoS One ; 11(5): e0156335, 2016.
Article in English | MEDLINE | ID: mdl-27227675

ABSTRACT

Identifying the characteristics of light interception and utilization is of great significance for improving the potential photosynthetic activity of plants. The present research investigates the differences in absorbing and converting photosynthetically active radiation (PAR) among various cotton cultivars. Field experiments were conducted in 2012, 2013 and 2014 in Anyang, Henan, China. Ten cultivars with different maturity and plant architectures were planted at a density of 60,000 plants ha-1 in randomized blocks, with three replicates. The spatial distribution of light in canopy was measured and quantified with a geo-statistical method, according to which the cumulative amount of intercepted radiation was calculated by Simpson 3/8 rules. Finally, light interception was analyzed in association with the biomass accumulation of different cultivars. The key results were: (1) late-maturing varieties with an incompact plant architecture captured more solar radiation throughout the whole growth period than middle varieties with columnar architecture and even more than early varieties with compact architecture, and they produced more biomass; (2) the highest PAR interception ratio and the maximum biomass accumulation rate occurred during the blossoming and boll-forming stage, when leaf area index (LAI) reached its peak; (3) the distribution within the canopy presented a significant spatial heterogeneity, and at late growing stage, the PAR was mainly intercepted by upper canopies in incompact-type plant communities, but was more homogeneous in columnar-type plants; however, the majority of radiation was transmitted through the canopy in compact-type colonies; (4) there was not a consistent variation relationship between the cumulative intercepted PAR (iPAR) and biomass among these cultivars over the three years of the study. Based on these results, we attempted to clarify the distinction in light spatial distribution within different canopies and the patterns of PAR interception in diverse cotton cultivars with different hereditary characters, thereby providing a significant basis for researchers to select cultivars with appropriate growth period and optimal plant architecture for improvement of light interception and utilization.


Subject(s)
Biomass , Gossypium/growth & development , Light , Plant Leaves/growth & development , Gossypium/metabolism , Gossypium/radiation effects , Photosynthesis , Plant Leaves/metabolism , Plant Leaves/radiation effects
11.
J Endod ; 42(3): 402-12, 2016 Mar.
Article in English | MEDLINE | ID: mdl-26709200

ABSTRACT

INTRODUCTION: Recombinant DNA-produced amelogenin protein was compared with calcium hydroxide in a study of immature apex closure conducted in 24 young mongrel dogs. METHODS: Root canals of maxillary and mandibular right premolars (n = 240) were instrumented and left open for 14 days. Canals were cleansed, irrigated, and split equally for treatment with recombinant mouse amelogenin (n = 120) or calcium hydroxide (n = 120). RESULTS: After 1, 3, and 6 months, the animals were sacrificed and the treated teeth recovered for histologic assessment and immunodetection of protein markers associated with odontogenic cells. After 1 month, amelogenin-treated canals revealed calcified tissue formed at the apical foramen and a pulp chamber containing soft connective tissue and hard tissue; amelogenin-treated canals assessed after 3- and 6-month intervals further included apical tissue functionally attached to bone by a periodontal ligament. In contrast, calcified apical tissue was poorly formed in the calcium hydroxide group, and soft connective tissue within the pulp chamber was not observed. CONCLUSIONS: The findings from this experimental strategy suggest recombinant amelogenin protein can signal cells to enhance apex formation in nonvital immature teeth and promote soft connective tissue regeneration.


Subject(s)
Amelogenin/pharmacology , Dental Pulp/drug effects , Dental Pulp/pathology , Regeneration/drug effects , Tooth Apex/drug effects , Tooth Apex/pathology , Animals , Apexification/methods , Calcium Hydroxide/pharmacology , Dental Pulp/growth & development , Dental Pulp Cavity/drug effects , Dental Pulp Cavity/growth & development , Dental Pulp Cavity/pathology , Dogs , Mice , Models, Animal , Odontoblasts/drug effects , Recombinant Proteins/pharmacology , Root Canal Filling Materials/pharmacology , Tooth Apex/growth & development , Tooth, Nonvital/pathology
12.
Biomaterials ; 61: 216-28, 2015 Aug.
Article in English | MEDLINE | ID: mdl-26004236

ABSTRACT

Tissue regeneration and development involves highly synchronized signals both between cells and with the extracellular environment. Biomaterials can be tuned to mimic specific biological signals and control cell response(s). As a result, these materials can be used as tools to elucidate cell signaling pathways and candidate molecules involved with cellular processes. In this work, we explore enamel-forming cells, ameloblasts, which have a limited regenerative capacity. By exposing undifferentiated cells to a self-assembling matrix bearing RGDS epitopes, we elicited a regenerative signal at will that subsequently led to the identification of thrombospondin 2 (TSP2), an extracellular matrix protein that has not been previously recognized as a key player in enamel development and regeneration. Targeted disruption of the thrombospondin 2 gene (Thbs2) resulted in enamel formation with a disordered architecture that was highly susceptible to wear compared to their wild-type counterparts. To test the regenerative capacity, we injected the bioactive matrix into the enamel organ and discovered that the enamel organic epithelial cells in TSP-null mice failed to polarize on the surface of the artificial matrix, greatly reducing integrin ß1 and Notch1 expression levels, which represent signaling pathways known to be associated with TSP2. These results suggest TSP2 plays an important role in regulating cell-matrix interactions during enamel formation. Exploiting the signaling pathways activated by biomaterials can provide insight into native signaling mechanisms crucial for tooth development and cell-based strategies for enamel regeneration.


Subject(s)
Ameloblasts/metabolism , Dental Enamel/physiology , Guided Tissue Regeneration/methods , Nanofibers/chemistry , Regeneration/physiology , Thrombospondins/metabolism , Ameloblasts/cytology , Ameloblasts/transplantation , Animals , Dental Enamel/cytology , Mice , Mice, Knockout , Thrombospondins/genetics
13.
J Biol Rhythms ; 27(3): 237-45, 2012 Jun.
Article in English | MEDLINE | ID: mdl-22653892

ABSTRACT

Fully mature enamel is about 98% mineral by weight. While mineral crystals appear very early during its formative phase, the newly secreted enamel is a soft gel-like matrix containing several enamel matrix proteins of which the most abundant is amelogenin (Amelx). Histological analysis of mineralized dental enamel reveals markings called cross-striations associated with daily increments of enamel formation, as evidenced by injections of labeling dyes at known time intervals. The daily incremental growth of enamel has led to the hypothesis that the circadian clock might be involved in the regulation of enamel development. To identify daily rhythms of clock genes and Amelx, we subjected murine ameloblast cells to serum synchronization to analyze the expression of the circadian transcription factors Per2 and Bmal1 by real-time PCR. Results indicate that these key genetic regulators of the circadian clock are expressed in synchronized murine ameloblast cell cultures and that their expression profile follows a circadian pattern with acrophase and bathyphase for both gene transcripts in antiphase. Immunohistological analysis confirms the protein expression of Bmal and Cry in enamel cells. Amelx expression in 2-day postnatal mouse molars dissected every 4 hours for a duration of 48 hours oscillated with an approximately 24-hour period, with a significant approximately 2-fold decrease in expression during the dark period compared to the light period. The expression of genes involved in bicarbonate production (Car2) and transport (Slc4a4), as well as in enamel matrix endocytosis (Lamp1), was greater during the dark period, indicating that ameloblasts express these proteins when Amelx expression is at the nadir. The human and mouse Amelx genes each contain a single nonconserved E-box element within 10 kb upstream of their respective transcription start sites. We also found that within 2 kb of the transcription start site of the human NFYA gene, which encodes a positive regulator of amelogenin, there is an E-box element that is conserved in rodents and other mammals. Moreover, we found that Nfya expression in serum-synchronized murine ameloblasts oscillated with a strong 24-hour rhythm. Taken together, our data support the hypothesis that the circadian clock temporally regulates enamel development.


Subject(s)
Amelogenin/biosynthesis , Circadian Rhythm , Dental Enamel/embryology , Gene Expression Regulation, Developmental , ARNTL Transcription Factors/metabolism , Ameloblasts/cytology , Amelogenin/genetics , Animals , Female , Humans , Mice , Models, Biological , Period Circadian Proteins/metabolism , Real-Time Polymerase Chain Reaction/methods , Species Specificity , Time Factors
14.
Cells Tissues Organs ; 194(2-4): 284-90, 2011.
Article in English | MEDLINE | ID: mdl-21540557

ABSTRACT

Two point mutations (T21I and P40T) within amelogenin have been identified from human DNA sequences in 2 instances of amelogenesis imperfecta. We studied the folding and self-assembly of recombinant amelogenin (rM180) compared to the T21I and P40T mutants analogs. At pH 5.8 and 25°C, rM180 and the P41T mutant existed as monomers, whereas the T21I mutant formed small oligomers. At pH 8 and 25°C, all of the amelogenin samples formed nanospheres with hydrodynamic radii (R(H)) of around 15-16 nm. Upon heating to 37°C, particles of P41T increased in size (R(H) = 18 nm). During thermal denaturation at pH 5.8, both of the mutant proteins refolded more slowly than the wild-type (WT) rM180. Variable temperature tryptophan fluorescence and dynamic light scattering studies showed that the WT transformed to a partially folded conformation upon heating and remained stable. Thermal denaturation and refolding studies indicated that the mutants were less stable and exhibit a greater ability to prematurely aggregate compared to the WT. Our data suggest that in the case of P41T, alterations in the self-assembly of amelogenin are a consequence of destabilization of the secondary structure, while in the case of T21I they are a consequence of change in the overall hydrophobicity at the N-terminal region. We propose that alterations in the assembly (i.e. premature aggregation) of mutant amelogenins may have a profound effect on intra- and extracellular processes such as amelogenin secretion, proteolysis, and its interactions with nonamelogenins as well as with the forming mineral.


Subject(s)
Amelogenin/chemistry , Amelogenin/genetics , Mutant Proteins/chemistry , Point Mutation/genetics , Protein Folding , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Amelogenin/metabolism , Amelogenin/ultrastructure , Animals , Circular Dichroism , Fluorescence , Humans , Hydrogen-Ion Concentration , Mice , Mutant Proteins/genetics , Mutant Proteins/metabolism , Protein Structure, Quaternary , Recombinant Proteins/metabolism , Recombinant Proteins/ultrastructure , Temperature
15.
Biomaterials ; 32(12): 3151-7, 2011 Apr.
Article in English | MEDLINE | ID: mdl-21295848

ABSTRACT

A biomimetic replacement for tooth enamel is urgently needed because dental caries is the most prevalent infectious disease to affect man. Here, design specifications for an enamel replacement material inspired by Nature are deployed for testing in an animal model. Using genetic engineering we created a simplified enamel protein matrix precursor where only one, rather than dozens of amelogenin isoforms, contributed to enamel formation. Enamel function and architecture were unaltered, but the balance between the competing materials properties of hardness and toughness was modulated. While the other amelogenin isoforms make a modest contribution to optimal biomechanical design, the enamel made with only one amelogenin isoform served as a functional substitute. Where enamel has been lost to caries or trauma a suitable biomimetic replacement material could be fabricated using only one amelogenin isoform, thereby simplifying the protein matrix parameters by one order of magnitude.


Subject(s)
Dental Enamel/metabolism , Genetic Engineering/methods , Mammals/genetics , Amelogenin/genetics , Amelogenin/metabolism , Animals , Dental Enamel/ultrastructure , Gene Expression Regulation , Gene Knock-In Techniques , Materials Testing , Mice , Physical Chromosome Mapping , RNA, Messenger/genetics , RNA, Messenger/metabolism
16.
J Biol Chem ; 285(52): 40593-603, 2010 Dec 24.
Article in English | MEDLINE | ID: mdl-20929860

ABSTRACT

Mutations in amelogenin sequence result in defective enamel, and the diverse group of genetically altered conditions is collectively known as amelogenesis imperfecta (AI). Despite numerous studies, the detailed molecular mechanism of defective enamel formation is still unknown. In this study, we have examined the biophysical properties of a recombinant murine amelogenin (rM180) and two point mutations identified from human DNA sequences in two cases of AI (T21I and P41T). At pH 5.8 and 25 °C, wild type (WT) rM180 and mutant P41T existed as monomers, and mutant T21I formed lower order oligomers. CD, dynamic light scattering, and fluorescence studies indicated that rM180 and P41T can be classified as a premolten globule-like subclass protein at 25 °C. Thermal denaturation and refolding monitored by CD ellipticity at 224 nm indicated the presence of a strong hysteresis in mutants compared with WT. Variable temperature tryptophan fluorescence and dynamic light scattering studies showed that WT transformed to a partially folded conformation upon heating and remained stable. The partially folded conformation formed by P41T, however, readily converted into a heterogeneous population of aggregates. T21I existed in an oligomeric state at room temperature and, upon heating, rapidly formed large aggregates over a very narrow temperature range. Thermal denaturation and refolding studies indicated that the mutants are less stable and exhibit poor refolding ability compared with WT rM180. Our results suggest that alterations in self-assembly of amelogenin are a consequence of destabilization of the intrinsic disorder. Therefore, we propose that, like a number of other human diseases, AI appears to be due to the destabilization of the secondary structure as a result of amelogenin mutations.


Subject(s)
Amelogenesis Imperfecta , Amelogenin/chemistry , Point Mutation , Protein Folding , Amelogenin/genetics , Amelogenin/metabolism , Humans , Protein Stability , Protein Structure, Secondary
17.
J Biol Chem ; 277(19): 17112-6, 2002 May 10.
Article in English | MEDLINE | ID: mdl-11877393

ABSTRACT

A hallmark of biological systems is a reliance on protein assemblies to perform complex functions. We have focused attention on mammalian enamel formation because it relies on a self-assembling protein complex to direct mineral habit. The principle protein of enamel is amelogenin, a 180-amino acid hydrophobic protein that self-assembles to form nanospheres. We have used independent technical methods, consisting of the yeast two-hybrid (Y2H) assay and surface plasmon resonance (SPR), to demonstrate the importance of amelogenin self-assembly domains. In addition, we have analyzed mutations in amelogenin observed in patients with amelogenesis imperfecta who demonstrate defects in enamel formation. Assessments of self-assembly of these mutant amelogenins by either SPR or Y2H assay yield concordant data. These data support the conclusion that the amelogenin amino-terminal self-assembly domain is essential to the creation of an enamel extracellular organic matrix capable of directing mineral formation. It also suggests that a pathway through which point mutations in the amelogenin protein can adversely impact on the formation of the enamel organ is by disturbing self-assembly of the organic matrix. These data support the utilization of the Y2H assay to search for protein interactions among extracellular matrix proteins that contribute to biomineralization and provide functional information on protein-protein and protein-mineral interactions.


Subject(s)
Amelogenesis Imperfecta/genetics , Dental Enamel Proteins/chemistry , Dental Enamel Proteins/metabolism , Genetic Linkage , X Chromosome , Amelogenin , Amino Acid Sequence , Animals , Dental Enamel/metabolism , Dental Enamel Proteins/genetics , Humans , Mice , Molecular Sequence Data , Mutation , Plasmids/metabolism , Protein Binding , Protein Structure, Tertiary , Surface Plasmon Resonance , Time Factors , Two-Hybrid System Techniques
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