Knuckles Research Articles

AGAMOUS regulates various target genes via cell cycle-coupled H3K27me3 dilution in floral meristems and stamens.

The MADS domain transcription factor AGAMOUS (AG) regulates floral meristem termination by preventing maintenance of the histone modification lysine 27 of histone H3 (H3K27me3) along the KNUCKLES (KNU) coding sequence. At 2 d after AG binding, cell division has diluted the repressive mark H3K27me3, allowing activation of KNU transcription prior to floral meristem termination. However, how many other downstream genes are temporally regulated by this intrinsic epigenetic timer and what their functions are remain unknown. Here, we identify direct AG targets regulated through cell cycle-coupled H3K27me3 dilution in Arabidopsis thaliana. Expression of the targets KNU, AT HOOK MOTIF NUCLEAR LOCALIZED PROTEIN18 (AHL18), and PLATZ10 occurred later in plants with longer H3K27me3-marked regions. We established a mathematical model to predict timing of gene expression and manipulated temporal gene expression using the H3K27me3-marked del region from the KNU coding sequence. Increasing the number of del copies delayed and reduced KNU expression in a polycomb repressive complex 2- and cell cycle-dependent manner. Furthermore, AHL18 was specifically expressed in stamens and caused developmental defects when misexpressed. Finally, AHL18 bound to genes important for stamen growth. Our results suggest that AG controls the timing of expression of various target genes via cell cycle-coupled dilution of H3K27me3 for proper floral meristem termination and stamen development.

Robust control of floral meristem determinacy by position-specific multifunctions of KNUCKLES

Floral organs are properly developed on the basis of timed floral meristem (FM) termination in Arabidopsis In this process, two known regulatory pathways are involved. The WUSCHEL (WUS)-CLAVATA3 (CLV3) feedback loop is vital for the spatial establishment and maintenance of the FM, while AGAMOUS (AG)-WUS transcriptional cascades temporally repress FM. At stage 6 of flower development, a C2H2-type zinc finger repressor that is a target of AG, KNUCKLES (KNU), directly represses the stem cell identity gene WUS in the organizing center for FM termination. However, how the robust FM activity is fully quenched within a limited time frame to secure carpel development is not fully understood. Here, we demonstrate that KNU directly binds to the CLV1 locus and the cis-regulatory element on CLV3 promoter and represses their expression during FM determinacy control. Furthermore, KNU physically interacts with WUS, and this interaction inhibits WUS from sustaining CLV3 in the central zone. The KNU-WUS interaction also interrupts the formation of WUS homodimers and WUS-HAIRYMERISTEM 1 heterodimers, both of which are required for FM maintenance. Overall, our findings describe a regulatory framework in which KNU plays a position-specific multifunctional role for the tightly controlled FM determinacy.

Expression of KNUCKLES in the Stem Cell Domain Is Required for Its Function in the Control of Floral Meristem Activity in Arabidopsis.

In the model plant Arabidopsis thaliana, the zinc-finger transcription factor KNUCKLES (KNU) plays an important role in the termination of floral meristem activity, a process that is crucial for preventing the overgrowth of flowers. The KNU gene is activated in floral meristems by the floral organ identity factor AGAMOUS (AG), and it has been shown that both AG and KNU act in floral meristem control by directly repressing the stem cell regulator WUSCHEL (WUS), which leads to a loss of stem cell activity. When we re-examined the expression pattern of KNU in floral meristems, we found that KNU is expressed throughout the center of floral meristems, which includes, but is considerably broader than the WUS expression domain. We therefore hypothesized that KNU may have additional functions in the control of floral meristem activity. To test this, we employed a gene perturbation approach and knocked down KNU activity at different times and in different domains of the floral meristem. In these experiments we found that early expression in the stem cell domain, which is characterized by the expression of the key meristem regulatory gene CLAVATA3 (CLV3), is crucial for the establishment of KNU expression. The results of additional genetic and molecular analyses suggest that KNU represses floral meristem activity to a large extent by acting on CLV3. Thus, KNU might need to suppress the expression of several meristem regulators to terminate floral meristem activity efficiently.

Knuckles.

Knuckles.

Integration of Transcriptional Repression and Polycomb-Mediated Silencing of WUSCHEL in Floral Meristems.

Arabidopsis (Arabidopsis thaliana) floral meristems terminate after the carpel primordia arise. This is achieved through the temporal repression of WUSCHEL (WUS), which is essential for stem cell maintenance. At floral stage 6, WUS is repressed by KNUCKLES (KNU), a repressor directly activated by AGAMOUS. KNU was suggested to repress WUS through histone deacetylation; however, how the changes in the chromatin state of WUS are initiated and maintained to terminate the floral meristem remains elusive. Here, we show that KNU integrates initial transcriptional repression with polycomb-mediated stable silencing of WUS After KNU is induced, it binds to the WUS promoter and causes eviction of SPLAYED, which is a known activator of WUS and can oppose polycomb repression. KNU also physically interacts with FERTILIZATION-INDEPENDENT ENDOSPERM, a key polycomb repressive complex2 component, and mediates the subsequent deposition of the repressive histone H3 lysine 27 trimethylation for stable silencing of WUS This multi-step silencing of WUS leads to the termination of floral stem cells, ensuring proper carpel development. Thus, our work describes a detailed mechanism for heritable floral stem cell termination in a precise spatiotemporal manner.

Induced ovule-to-flower switch by interfering with SlIMA activity in tomato

ABSTRACTThe INHIBITOR OF MERISTEM ACTIVITY in tomato (SlIMA) and MINI ZINC FINGER 2 in Arabidopsis (AtMIF2), two members of the MINI ZINC FINGER family (MIF), are involved in the regulation of flower and ovule development. MIF proteins possess a unique non-canonical zinc-finger domain that confers the capacity to interact with other protein partners. The characterization of SlIMA and AtMIF2 gain- and loss-of-function transgenic lines in Solanum lycopersicum and Arabidopsis thaliana respectively, allowed the demonstration of their conserved functional role in the termination of floral stem cell maintenance. During early floral development, the expression of SlIMA and AtMIF2 is induced by the MADS-Box transcription factor AGAMOUS (AG). Then, SlIMA or AtMIF2 protein recruits the C2H2 zinc finger KNUCKLES (KNU), in a transcriptional repressor complex together with TOPLESS (TPL) and HISTONE DEACETYLASE19 (HDA19). This complex binds to the WUSCHEL (WUS) locus leading to its repression. To further characterize the role of these interactions in flower development, we have investigated the effects of a dominant negative form of SlIMA, SlIMAch that leads to spectacular phenotypes, including ovule conversion into a floral meristem.

Why do your knuckles crack? Engineers say they finally figured it out

Why do your knuckles crack? Engineers say they finally figured it out

At-MINI ZINC FINGER2 and Sl-INHIBITOR OF MERISTEM ACTIVITY, a Conserved Missing Link in the Regulation of Floral Meristem Termination in Arabidopsis and Tomato.

In angiosperms, the gynoecium is the last structure to develop within the flower due to the determinate fate of floral meristem (FM) stem cells. The maintenance of stem cell activity before its arrest at the stage called FM termination affects the number of carpels that develop. The necessary inhibition at this stage of WUSCHEL (WUS), which is responsible for stem cell maintenance, involves a two-step mechanism. Direct repression mediated by the MADS domain transcription factor AGAMOUS (AG), followed by indirect repression requiring the C2H2 zinc-finger protein KNUCKLES (KNU), allow for the complete termination of floral stem cell activity. Here, we show that Arabidopsis thaliana MINI ZINC FINGER2 (AtMIF2) and its homolog in tomato (Solanum lycopersicum), INHIBITOR OF MERISTEM ACTIVITY (SlIMA), participate in the FM termination process by functioning as adaptor proteins. AtMIF2 and SlIMA recruit AtKNU and SlKNU, respectively, to form a transcriptional repressor complex together with TOPLESS and HISTONE DEACETYLASE19. AtMIF2 and SlIMA bind to the WUS and SlWUS loci in the respective plants, leading to their repression. These results provide important insights into the molecular mechanisms governing (FM) termination and highlight the essential role of AtMIF2/SlIMA during this developmental step, which determines carpel number and therefore fruit size.

Stop Cracking Your Knuckles!

Stop Cracking Your Knuckles!

Finite Element Simulation of Knuckle and Strut Arm Column Assembly for Automotive Steering System

Steering knuckle is one of the important parts in the vehicle steering system. Under certain operating conditions, the knuckles reliability threatens the safety of people and vehicles directly. At emergency braking condition, mostly strut arm and steering arm have maximum deflection in steering knuckle when it is subjected to various load cases. In this work, finite element analysis of the spheroid graphite (SG) iron strut arm of steering knuckle with strut mount assembly was carried out to predict its deflection under static load. The analysis result was compared with that of the experimental results to put forward directions to optimize the shape and material selection.

Timing Mechanism Dependent on Cell Division Is Invoked by Polycomb Eviction in Plant Stem Cells

Plant floral stem cells divide a limited number of times before they stop and terminally differentiate, but the mechanisms that control this timing remain unclear. The precise temporal induction of the Arabidopsis zinc finger repressor KNUCKLES (KNU) is essential for the coordinated growth and differentiation of floral stem cells. We identify an epigenetic mechanism in which the floral homeotic protein AGAMOUS (AG) induces KNU at ~2 days of delay. AG binding sites colocalize with a Polycomb response element in the KNU upstream region. AG binding to the KNU promoter causes the eviction of the Polycomb group proteins from the locus, leading to cell division-dependent induction. These analyses demonstrate that floral stem cells measure developmental timing by a division-dependent epigenetic timer triggered by Polycomb eviction.

Fancy Limes at a Sponging House and The Biographical Fallacies Found in Migration’s Anxious Knuckles

Fancy Limes at a Sponging House and The Biographical Fallacies Found in Migration’s Anxious Knuckles

Investigation on researching and clinical application of special gloves for medical needle

Objective To investigate the researching and clinical application of special gloves for medical needle,in order to solve the problem of adhesive plaster' s sticking to gloves in medical personnel during puncture operation and reduce the risk of needle stick injury and prevention of iatrogenic infection.Methods Key technology is the structure of the glove's index fmger,middle finger and thumb for technological reform,that is,on the basis of the original gloves,at the distal of left and right hand index fingers,two middle fingers' knuckles and distal thumb fingertip outer plane,we add a layer of material of waterproof adhesive piaster,and then investigate its application conditions in medical institutions.Results Medical personnel in the region accepted and welcomed this type of gloves ; it improved patients' satisfaction degree; after using this type of glove,the needle injury cases reduced from 36 to 12,it also improved the productivity,and made the expenditure for blood-spreading diseases decrease from 31 680 RMB to 10560 RMB,reducing hospital expenditure and increased social benefits.Conclusion It is necessary to study this type of gloves,and investigate its applications to solve the problems of adhesive plaster sticking to glove,reduce the risk of needle sticking injury,decrease the medical expenditure and improve the efficiency of medical personnel.It has been recognized by the medical staff,worthy of promotion.This glove has got the certificate of utility model patent on May 18,2011 issued by The People's Republic of China State Intellectual Property Bureau,with patent number ZL 2010 2 0551435.2 Key words: Special gloves for medical needle; Development; Clinical application; Investigation

Dermatomyositis coupled with extensive calcinosis: a case report

A case of dermatomyositis coupled with extensive calcinosis is reported. A 25-year-old male presented with a 10-year history of dermatomyositis. Two years prior to the presentation, subcutaneous calcinosis emerged on the waist, and rapidly involved the trunk and limbs with limited mobility. On admission,the symptom of amyasthenia was remarkable with extensive prunosus and edematous macules all over the body surface, many large firm irregularly-sized subcutaneous masses on the trunk and limbs, limited mobility of knuckles in the flexed posture. There was also a marked increase of serum creatase. After treatment with prednisone, methotrexate, hydroxychloroquine for 4 weeks, an evident improvement was observed in the lesions and amyasthenia along with a reduction in serum creatase level. The use of diltiazem was attempted to manage calcinosis in this patient, who was still under follow up till the time of this writing. Key words: Dermatomyositis; Calcinosis; Diltiazem

A timing mechanism for stem cell maintenance and differentiation in the Arabidopsis floral meristem

Developmental regulation of the floral meristem ensures that plants of the same species have similarly sized flowers with a fixed number of floral organs. The maintenance of stem cells in the floral meristem is terminated after the production of a fixed number of floral organ primordia. Precise repression of the Arabidopsis thaliana homeobox gene WUSCHEL (WUS) by the floral homeotic protein AGAMOUS (AG) plays a major part in this process. Here we show that KNUCKLES (KNU) mediates the repression of WUS in floral meristem determinacy control. AG directly induces the transcription of KNU, which encodes a C2H2-type zinc finger protein with a conserved transcriptional repression motif. In turn, KNU represses WUS transcription to abolish stem cell activity. We also show that the timing of KNU induction is key in balancing proliferation and differentiation in flower development. Delayed KNU expression results in an indeterminate meristem, whereas ectopic KNU expression prematurely terminates the floral meristem. Furthermore, KNU induction by AG is preceded by changes in repressive histone modification at the KNU locus, which occurs in an AG-dependent manner. This study provides a mechanistic link between transcriptional feedback and epigenetic regulation in plant stem cell proliferation.

A terminator of floral stem cells: Figure 1.

Normal flower development requires the termination of stem cell activities in the floral meristem. The floral regulator AGAMOUS (AG) is necessary for this termination and represses the expression of the stem cell determinant WUSCHEL (WUS), but the repression mechanism was not clear. A recent study by Sun and colleagues (pp. 1791-1804) in this issue of Genes & Development has identified a direct target of AG, KNUCKLES (KNU), which encodes a transcriptional repressor of WUS, providing a key missing link in floral meristem determinacy.

The ovipositional behaviour of the endemic whistling lizard Calotes liolepis Boulenger, 1885 (Reptilia: Agamidae) in the Knuckles forest region of Sri Lanka

A mature female Calotes liolepis was observed laying eggs on the ground in Manigala in the Knuckles Forest Region of Sri Lanka. This is the first completely described observation of the ovipositioning as well as the captive egg hatching of Calotes liolepis . This ovipositional behaviour consisted of the digging of the nest cavity, oviposition, scraping of the soil to bury the eggs, filling of the spaces between the eggs, compression of the soil, and camouflage of the nest. The sizes of three eggs were increased during incubation: day 1 mean = 17.5 mm × 9.2 mm (length × width), and after 70-71 days mean = 21.7 mm × 14.4 mm. Three hatchlings were emerged (mean snout-to-vent length = 29.9 mm; tail length = 58.2 mm; head length = 10.2 mm. Immediate conservation measures are needed for this endemic and threatened lizard, and the observations related to its egg-laying could be useful in planning and implementing suitable conservation methods.

Don't Knock These Knuckles

Don't Knock These Knuckles

Of Whale Knuckles and Placental Trees

The Rise of Placental Mammals . Origins and Relationships of the Major Extant Clades. Kenneth D. Rose and J. David Archibald, Eds. . Johns Hopkins University Press, Baltimore, MD, 2005. 271 pp. $95, £63.50. ISBN 0-8018-8022-X. Focusing on anatomical evidence but not ignoring findings from molecular studies, the contributors discuss our current understanding of the initial radiation and ordinal relationships of placental mammals.

KNUCKLES (KNU) encodes a C2H2 zinc-finger protein that regulates development of basal pattern elements of the Arabidopsis gynoecium.

Flowers of the parthenocarpic knuckles mutant are conditionally male sterile and contain ectopic stamens and carpels that originate from placental tissue within developing gynoecia. The mutation was mapped to a 123 Kb interval on chromosome 5 using molecular markers. All aspects of the knuckles phenotype could be complemented by a genomic fragment from the region which contained the annotated MAC12.2 gene. A guanine to adenine transition within a predicted C2H2 zinc finger-encoding region of MAC12.2 causes the second of two critical zinc-binding cysteine residues to be replaced by a tyrosine. Transgenic plants in which translational fusions of the GUS reporter to KNUCKLES were driven by the presumptive KNUCKLES promoter indicate that the gene is expressed first in developing carpel primordia, and later in stamens and ovules of flower buds. In situ hybridization experiments showed a broader pattern of transcript localization, suggesting that post-transcriptional regulatory mechanisms may limit KNUCKLES protein accumulation and localization. Based on genetic evidence and the presence of a carboxy-terminal motif demonstrated by others to function as an active repression domain, we propose that KNUCKLES might function as a transcriptional repressor of cellular proliferation that regulates floral determinacy and relative size of basal pattern elements along the proximo-distal axis of the developing Arabidopsis gynoecium.