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Abstract Acromioclavicular joint dislocations often occur in athletic, young patients after blunt force to the shoulder. Several static and dynamic operative procedures with or without primary ligament replacement have been described. Between February 2003 and March 2009 we treated 313 patients suffering from Rockwood III-V lesions of the AC joint with an AC-hook plate. 225 (72%) of these patients could be followed up. Mean operation time was 42 minutes in the conventional group and 47 minutes in the minimal invasive group. The postoperative pain on a scale from 1 to 10 (VAS-scale) was rated 2.7 in the conventional group and 2.2 in the minimal invasive group. Taft score showed very good and good results in 189 patients (84%). Constant score showed an average of 92.4 of 100 possible points with 89% excellent and good results and 11% satisfying results. All patients had some degree of pain or discomfort with the hookplate in place. These symptoms were relieved after removal of the plate. The overall complication rate was 10.6%. There were 6 superficial soft tissue infections, 1 fracture of the acromion, 7 redislocations after removal of the hook-plate. We observed 4 broken hooks which could be removed at the time of plate removal, 4 seromas and 2 cases of lateral clavicle bone infection, which required early removal of the plate. We can conclude that clavicle hook plate is a convenient device for the surgical treatment of Rockwood Grade III-V dislocations, giving good mid-term results with a low overall complication rate compared to the literature. Early functional therapy is possible and can avoid limitations in postoperative shoulder function.

Abstract The cut-out of the sliding screw is one of the most common complications in the treatment of intertrochanteric fractures. The reasons for the cut-out are: a suboptimal position of the hip-screw in the femoral head, the type of fracture and poor bone quality. The aim of this study was to reproduce the cut-out event biomechanically and to evaluate the possible prevention of this event by the use of a biopolymer augmentation of the hip screw. Concerning the density and compression force of osteoporotic femoral bone polyurethane foam according to the terms of the Association for Standard Testing Material (ASTMF 1839-97) was used as test material. The polyurethane foam Lumoltan 200 with a compression force of 3.3 Mpa and a density of 0.192 g/cm3 was used to reproduce the osteoporotic bone of the femoral fragment (density 12 lbm/ft3). A cylinder of 50 mm of length and 50 mm of width was produced by a rotary splint raising procedure with planar contact. The axial load of the system was performed by a hydraulic force cylinder of a universal test machine type Zwick 1455, Ulm, Germany. The CCD-angle of the used TGN-System was preset at 130 degrees. The migration pattern of the hip screw in the polyurethane foam was measured and expressed as a curve of the distance in millimeter [mm] against the applied load in Newton [N] up to the cut-out point. During the tests the implants reached a critical changing point from stable to unstable with an increased load progression of steps of 50 Newton. This unstable point was characterized by an increased migration speed in millimeters and higher descending gradient in the migration curve. This peak of the migration curve served as an indicator for the change of the hip screw position in the simulated bone material. The applied load in the non-augmented implant showed that in this group for a density degree of 12 (0,192 g/cm3) the mean force at the failure point was 1431 Newton (± 52 Newton). In the augmented implant we found that the mean force at the failure point was 1987 Newton (± 84 Newton). This difference was statistically significant. In conclusion, the bone density is a significant factor for the stability of the hip screw implant. The osteosynthesis with screws in material with low density increases the chance for cut-out. A biopolymer augmented hip screw could significantly improve the stability of the fixation. The use of augmentation with a fast hardening bone replacement material containing polymer-ceramic changes the point of failure under axial load in the osteoporotic bone model and could significantly improve the failure point. Our study results indicate, that a decrease of failure in terms of cut-out can be achieved with polymer augmentation of hip screws in osteoporotic bones.

Insulators are cis -regulatory elements that separate transcriptional units, whereas silencers are elements that repress transcription regardless of their position. In plants, these elements remain largely uncharacterized. Here, we use the massively parallel reporter assay Plant STARR-seq with short fragments of eight large insulators to identify more than 100 fragments that block enhancer activity. The short fragments can be combined to generate more powerful insulators that abolish the capacity of the strong viral 35S enhancer to activate the 35S minimal promoter. Unexpectedly, when tested upstream of weak enhancers, these fragments act as silencers and repress transcription. Thus, these elements are capable of both insulating or repressing transcription dependent upon regulatory context. We validate our findings in stable transgenic Arabidopsis , maize, and rice plants. The short elements identified here should be useful building blocks for plant biotechnology efforts., Competing Interests: Competing interests T.J., J.T.C., and C.Q. have filed a patent application related to this work through the University of Washington. The remaining authors declare no competing interests.

The precise onset of flowering is crucial to ensure successful plant reproduction. The gene FLOWERING LOCUS T ( FT ) encodes florigen, a mobile signal produced in leaves that initiates flowering at the shoot apical meristem. In response to seasonal changes, FT is induced in phloem companion cells located in distal leaf regions. Thus far, a detailed molecular characterization of the FT -expressing cells has been lacking. Here, we used bulk nuclei RNA-seq and single nuclei RNA (snRNA)-seq to investigate gene expression in FT -expressing cells and other phloem companion cells. Our bulk nuclei RNA-seq demonstrated that FT -expressing cells in cotyledons and in true leaves differed transcriptionally. Within the true leaves, our snRNA-seq analysis revealed that companion cells with high FT expression form a unique cluster in which many genes involved in ATP biosynthesis are highly upregulated. The cluster also expresses other genes encoding small proteins, including the flowering and stem growth inducer FPF1-LIKE PROTEIN 1 (FLP1) and the anti-florigen BROTHER OF FT AND TFL1 (BFT). In addition, we found that the promoters of FT and the genes co-expressed with FT in the cluster were enriched for the consensus binding motifs of NITRATE-INDUCIBLE GARP-TYPE TRANSCRIPTIONAL REPRESSOR 1 (NIGT1). Overexpression of the paralogous NIGT1.2 and NIGT1.4 repressed FT expression and significantly delayed flowering under nitrogen-rich conditions, consistent with NIGT1s acting as nitrogen-dependent FT repressors. Taken together, our results demonstrate that major FT -expressing cells show a distinct expression profile that suggests that these cells may produce multiple systemic signals to regulate plant growth and development., Competing Interests: Competing Interest Statement: The authors declare no competing interest.

Many of the most highly conserved elements in the human genome are "poison exons," alternatively spliced exons that contain premature termination codons and permit post-transcriptional regulation of mRNA abundance through induction of nonsense-mediated mRNA decay (NMD). Poison exons are widely assumed to be highly conserved due to their presumed importance for organismal fitness, but this functional importance has never been tested in the context of a whole organism. Here, we report that a poison exon in Smndc1 is conserved across mammals and plants and plays a molecular autoregulatory function in both kingdoms. We generated mouse and A. thaliana models lacking this poison exon to find its loss leads to deregulation of SMNDC1 protein levels, pervasive alterations in mRNA processing, and organismal size restriction. Together, these models demonstrate the importance of poison exons for both molecular and organismal phenotypes that likely explain their extraordinary conservation., Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: R.K.B. is a founder and scientific advisor of Codify Therapeutics and Synthesize Bio and holds equity in both companies. R.K.B. has received research funding from Codify Therapeutics unrelated to the current work., (Copyright: © 2024 Belleville et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Since their initial discovery in maize, transposable elements (TEs) have emerged as being integral to the evolution of maize, accounting for 80% of its genome. However, the repetitive nature of TEs has hindered our understanding of their regulatory potential. Here, we demonstrate that long-read chromatin fiber sequencing (Fiber-seq) permits the comprehensive annotation of the regulatory potential of maize TEs. We uncover that only 94 LTR retrotransposons contain the functional epigenetic architecture required for mobilization within maize leaves. This epigenetic architecture degenerates with evolutionary age, resulting in solo TE enhancers being preferentially marked by simultaneous hyper-CpG methylation and chromatin accessibility, an architecture markedly divergent from canonical enhancers. We find that TEs shape maize gene regulation by creating novel promoters within the TE itself as well as through TE-mediated gene amplification. Lastly, we uncover a pervasive epigenetic code directing TEs to specific loci, including that locus that sparked McClintock's discovery of TEs., Competing Interests: Competing interests. A.B.S. is a co-inventor on a patent relating to the Fiber-seq method (US17/995,058).

The 3' end of a gene, often called a terminator, modulates mRNA stability, localization, translation, and polyadenylation. Here, we adapted Plant STARR-seq, a massively parallel reporter assay, to measure the activity of over 50,000 terminators from the plants Arabidopsis thaliana and Zea mays. We characterize thousands of plant terminators, including many that outperform bacterial terminators commonly used in plants. Terminator activity is species-specific, differing in tobacco leaf and maize protoplast assays. While recapitulating known biology, our results reveal the relative contributions of polyadenylation motifs to terminator strength. We built a computational model to predict terminator strength and used it to conduct in silico evolution that generated optimized synthetic terminators. Additionally, we discover alternative polyadenylation sites across tens of thousands of terminators; however, the strongest terminators tend to have a dominant cleavage site. Our results establish features of plant terminator function and identify strong naturally occurring and synthetic terminators., (© 2024. The Author(s).)

Enhancers are cis-regulatory elements that shape gene expression in response to numerous developmental and environmental cues. In animals, several models have been proposed to explain how enhancers integrate the activity of multiple transcription factors. However, it remains largely unclear how plant enhancers integrate transcription factor activity. Here, we use Plant STARR-seq to characterize 3 light-responsive plant enhancers-AB80, Cab-1, and rbcS-E9-derived from genes associated with photosynthesis. Saturation mutagenesis revealed mutations, many of which clustered in short regions, that strongly reduced enhancer activity in the light, in the dark, or in both conditions. When tested in the light, these mutation-sensitive regions did not function on their own; rather, cooperative interactions with other such regions were required for full activity. Epistatic interactions occurred between mutations in adjacent mutation-sensitive regions, and the spacing and order of mutation-sensitive regions in synthetic enhancers affected enhancer activity. In contrast, when tested in the dark, mutation-sensitive regions acted independently and additively in conferring enhancer activity. Taken together, this work demonstrates that plant enhancers show evidence for both cooperative and additive interactions among their functional elements. This knowledge can be harnessed to design strong, condition-specific synthetic enhancers., Competing Interests: Conflict of interest statement. None declared., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society of Plant Biologists.)

The rich diversity of angiosperms, both the planet's dominant flora and the cornerstone of agriculture, is integrally intertwined with a distinctive evolutionary history. Here, we explore the interplay between angiosperm genome organization and botanical diversity, empowered by genomic approaches ranging from genetic linkage mapping to analysis of gene regulation. Commonality in the genetic hardware of plants has enabled robust comparative genomics that has provided a broad picture of angiosperm evolution and implicated both general processes and specific elements in contributing to botanical diversity. We argue that the hardware of plant genomes-both in content and in dynamics-has been shaped by selection for rather substantial differences in gene regulation between plants and animals such as maize and human, organisms of comparable genome size and gene number. Their distinctive genome content and dynamics may reflect in part the indeterminate development of plants that puts strikingly different demands on gene regulation than in animals. Repeated polyploidization of plant genomes and multiplication of individual genes together with extensive rearrangement and differential retention provide rich raw material for selection of morphological and/or physiological variations conferring fitness in specific niches, whether natural or artificial. These findings exemplify the burgeoning information available to employ in increasing knowledge of plant biology and in modifying selected plants to better meet human needs., Competing Interests: Conflict of interest statement. None declared., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society of Plant Biologists. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Isogenic individuals can display seemingly stochastic phenotypic differences, limiting the accuracy of genotype-to-phenotype predictions. The extent of this phenotypic variation depends in part on genetic background, raising questions about the genes involved in controlling stochastic phenotypic variation. Focusing on early seedling traits in Arabidopsis thaliana, we found that hypomorphs of the cuticle-related gene LIPID TRANSFER PROTEIN 2 (LTP2) greatly increased variation in seedling phenotypes, including hypocotyl length, gravitropism and cuticle permeability. Many ltp2 hypocotyls were significantly shorter than wild-type hypocotyls while others resembled the wild-type. Differences in epidermal properties and gene expression between ltp2 seedlings with long and short hypocotyls suggest a loss of cuticle integrity as the primary determinant of the observed phenotypic variation. We identified environmental conditions that reveal or mask the increased variation in ltp2 hypomorphs and found that increased expression of its closest paralog LTP1 is necessary for ltp2 phenotypes. Our results illustrate how decreased expression of a single gene can generate starkly increased phenotypic variation in isogenic individuals in response to an environmental challenge., (© 2023 The Authors New Phytologist © 2023 New Phytologist Foundation.)

Embryonic development is remarkably robust, but temperature stress can degrade its ability to generate animals with invariant anatomy. Phenotypes associated with environmental stress suggest that some cell types are more sensitive to stress than others, but the basis of this sensitivity is unknown. Here, we characterize hundreds of individual zebrafish embryos under temperature stress using whole-animal single-cell RNA sequencing (RNA-seq) to identify cell types and molecular programs driving phenotypic variability. We find that temperature perturbs the normal proportions and gene expression programs of numerous cell types and also introduces asynchrony in developmental timing. The notochord is particularly sensitive to temperature, which we map to a specialized cell type: sheath cells. These cells accumulate misfolded protein at elevated temperature, leading to a cascading structural failure of the notochord and anatomic defects. Our study demonstrates that whole-animal single-cell RNA-seq can identify mechanisms for developmental robustness and pinpoint cell types that constitute key failure points., Competing Interests: Declaration of interests C.T. is a scientific advisory board (SAB) member, consultant, and/or co-founder of Algen Biotechnologies, Altius Therapeutics, and Scale Biosciences. J.S. is an SAB member, consultant, and/or co-founder of Cajal Neuroscience, Guardant Health, Maze Therapeutics, Camp4 Therapeutics, Phase Genomics, Adaptive Biotechnologies, and Scale Biosciences., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Understanding plant gene regulation has been a priority for generations of plant scientists. However, due to its complex nature, the regulatory code governing plant gene expression has yet to be deciphered comprehensively. Recently developed methods-often relying on next-generation sequencing technology and state-of-the-art computational approaches-have started to further our understanding of the gene regulatory logic used by plants. In this review, we discuss these methods and the insights into the regulatory code of plants that they can yield., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

The transfection of maize mesophyll cells often involves digesting the plant cell walls to create protoplasts and then inserting DNA via electroporation or polyethylene glycol (PEG). Previous methods were developed to produce tens of thousands of transfected protoplasts at once. Here, we describe a straightforward method to isolate and transfect millions of leaf mesophyll protoplasts in maize (Zea mays L.). This streamlined process removes certain common protoplasting steps, such as washing in W5. Additionally, steps such as centrifugation, PEG-mediated transfection, and incubation have been modified to work with a greater number of protoplasts. The ability to express large libraries of plasmid constructs enables genome-scale experiments, such as massively parallel reporter assays in maize.

Background: The genetic information contained in the genome of an organism is organized in genes and regulatory elements that control gene expression. The genomes of multiple plants species have already been sequenced and the gene repertory have been annotated, however, cis-regulatory elements remain less characterized, limiting our understanding of genome functionality. These elements act as open platforms for recruiting both positive- and negative-acting transcription factors, and as such, chromatin accessibility is an important signature for their identification., Results: In this work we developed a transgenic INTACT [isolation of nuclei tagged in specific cell types] system in tetraploid wheat for nuclei purifications. Then, we combined the INTACT system together with the assay for transposase-accessible chromatin with sequencing [ATAC-seq] to identify open chromatin regions in wheat root tip samples. Our ATAC-seq results showed a large enrichment of open chromatin regions in intergenic and promoter regions, which is expected for regulatory elements and that is similar to ATAC-seq results obtained in other plant species. In addition, root ATAC-seq peaks showed a significant overlap with a previously published ATAC-seq data from wheat leaf protoplast, indicating a high reproducibility between the two experiments and a large overlap between open chromatin regions in root and leaf tissues. Importantly, we observed overlap between ATAC-seq peaks and cis-regulatory elements that have been functionally validated in wheat, and a good correlation between normalized accessibility and gene expression levels., Conclusions: We have developed and validated an INTACT system in tetraploid wheat that allows rapid and high-quality nuclei purification from root tips. Those nuclei were successfully used to performed ATAC-seq experiments that revealed open chromatin regions in the wheat genome that will be useful to identify cis-regulatory elements. The INTACT system presented here will facilitate the development of ATAC-seq datasets in other tissues, growth stages, and under different growing conditions to generate a more complete landscape of the accessible DNA regions in the wheat genome., (© 2023. The Author(s).)

The genes that encode ribosomal RNAs are present in several hundred copies in most eukaryotes. These vast arrays of repetitive ribosomal DNA (rDNA) have been implicated not just in ribosome biogenesis, but also aging, cancer, genome stability, and global gene expression. rDNA copy number is highly variable among and within species; this variability is thought to associate with traits relevant to human health and disease. Here we investigate the phenotypic consequences of multicellular life at the lower bounds of rDNA copy number. We use the model Caenorhabditis elegans, which has previously been found to complete embryogenesis using only maternally provided ribosomes. We find that individuals with rDNA copy number reduced to ∼5% of wild type are capable of further development with variable penetrance. Such individuals are sterile and exhibit severe morphological defects, particularly in post-embryonically dividing tissues such as germline and vulva. Developmental completion and fertility are supported by an rDNA copy number ∼10% of wild type, with substantially delayed development. Worms with rDNA copy number reduced to ∼33% of wild type display a subtle developmental timing defect that was absent in worms with higher copy numbers. Our results support the hypothesis that rDNA requirements vary across tissues and indicate that the minimum rDNA copy number for fertile adulthood is substantially less than the lowest naturally observed total copy number. The phenotype of individuals with severely reduced rDNA copy number is highly variable in penetrance and presentation, highlighting the need for continued investigation into the biological consequences of rDNA copy number variation., Competing Interests: Conflicts of interest The authors declare no conflict of interest., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Genetics Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Timely completion of genome replication is a prerequisite for mitosis, genome integrity, and cell survival. A challenge to this timely completion comes from the need to replicate the hundreds of untranscribed copies of rDNA that organisms maintain in addition to the copies required for ribosome biogenesis. Replication of these rDNA arrays is relegated to late S phase despite their large size, repetitive nature, and essentiality. Here, we show that, in Saccharomyces cerevisiae, reducing the number of rDNA repeats leads to early rDNA replication, which results in delaying replication elsewhere in the genome. Moreover, cells with early-replicating rDNA arrays and delayed genome-wide replication aberrantly release the mitotic phosphatase Cdc14 from the nucleolus and enter anaphase prematurely. We propose that rDNA copy number determines the replication time of the rDNA locus and that the release of Cdc14 upon completion of rDNA replication is a signal for cell cycle progression., Competing Interests: Declaration of interests C.Q. is a Cell Reports Advisory Board member, Ecology and Evolution., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)