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Mitochondrial function [e.g., respiration, respiratory coupling ratio (RCR) and reactive oxygen species (ROS) production] is negatively impacted by high temperatures, and may limit fish cardiac performance and upper thermal tolerance. Nevertheless, direct evidence of this is lacking, and testing at different levels of organization will be critical to establishing whether this construct has validity. We acclimated salmon to 10 and 20°C for &amp;gt;2 weeks and then: used strips from half of the ventricle to simultaneously measure muscle work and O2 consumption (and thus efficiency) at 20 and 26oC; and the other half to obtain myocardial homogenates and isolated mitochondria so that mitochondrial functional parameters could be measured at the same temperatures. State 4 respiration for isolated mitochondria and State 2 respiration for homogenates, and absolute ROS production during these states, were positively correlated with resting muscle O2 consumption in fish acclimated to both temperatures. In contrast, we were unable to consistently identify relationships between mitochondrial parameters and O2 consumption/efficiency while the muscle was working. For example, the P:O ratio (Complex I) and RCR (Complex I+II) for these preparations were not related to the absolute efficiency of shortening work. Further, while total CI+CII ROS production was negatively correlated with the efficiency of shortening work in 20oC acclimated salmon, this relationship was not evident for 10oC acclimated fish. These results: are consistent with previous data showing that warm acclimation improves cardiac mitochondrial function at high temperatures; but only provide a tenuous link between heart and mitochondrial performance in salmon under such conditions.

SEB Conference 2022

Are Cardiac Muscle Performance and Mitochondrial Function Related in Atlantic Salmon (Salmo salar) at High Temperatures?

Mitochondrial function [e.g., respiration, respiratory coupling ratio (RCR) and reactive oxygen species (ROS) production] is negatively impacted by high temperatures, and may limit fish cardiac performance and upper thermal tolerance. Nevertheless, direct evidence of this is lacking, and testing at different levels of organization will be critical to establishing whether this construct has validity. We acclimated salmon to 10 and 20°C for &gt;2 weeks and then: used strips from half of the ventricle to simultaneously measure muscle work and O2 consumption (and thus efficiency) at 20 and 26oC; and the other half to obtain myocardial homogenates and isolated mitochondria so that mitochondrial functional parameters could be measured at the same temperatures. State 4 respiration for isolated mitochondria and State 2 respiration for homogenates, and absolute ROS production during these states, were positively correlated with resting muscle O2 consumption in fish acclimated to both temperatures. In contrast, we were unable to consistently identify relationships between mitochondrial parameters and O2 consumption/efficiency while the muscle was working. For example, the P:O ratio (Complex I) and RCR (Complex I+II) for these preparations were not related to the absolute efficiency of shortening work. Further, while total CI+CII ROS production was negatively correlated with the efficiency of shortening work in 20oC acclimated salmon, this relationship was not evident for 10oC acclimated fish. These results: are consistent with previous data showing that warm acclimation improves cardiac mitochondrial function at high temperatures; but only provide a tenuous link between heart and mitochondrial performance in salmon under such conditions.

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2022 found over 1500 experimental biologists from across the globe converging on the third-largest French city near the Mediterranean coast and the fastest-growing city in the country over the past 25 years - Montpellier, France.

Many small vertebrates and arthropods are able to run with sticky feet, and must hence be able to rapidly switch between strong attachment and effortless detachment. The timescale over which contacts are formed and broken precludes chemical control. Instead, terrestrial climbing animals appear to rely on mechanical `switches’. Recently, strong evidence has emerged that at least some of these switches are universal: Whether adhesive pads are `hairy’, `smooth’, `wet’ or `dry’ pads, there appears to exist a linear relationship between the shear force, applied in parallel to the surface, and the adhesive force, normal to the surface, required to detach them. This ‘shear-sensitivity’ provides a rapid mechanical switch, enables larger animals to maintain approximately constant safety factors despite smaller surface-to-volume ratios, and has implications for optimal step coordination patterns during climbing on vertical and inverted substrates. The control of adhesion via shear forces is deeply integrated with the anatomy and locomotion of climbing animals, and involves both active neuro-muscular control, and rapid passive responses. The resulting dynamic adhesive systems are robust, reliable, versatile and nevertheless remarkably simple. They may thus serve as suitable inspiration for the design of climbing robots.

Biomechanics of controllable attachment in climbing animals

There is a wealth of evidence for the impact that early-life conditions, e.g., differences in growth rate, have on individuals’ later-life phenotype and performance. Mitochondria are believed to be a key player in this, as they not only fuel the cellular processes but are also involved in health and age-related disorders. Birds have surprisingly slow ageing rates and long lifespans for their body size and physiology. Recent work with avian models has highlighted the utility of determining mitochondrial function in their nucleated red blood cells (RBCs), which enables longitudinal sampling of mitochondria within individuals as they age. Here, we employ this new perspective to test, using zebra finches (Taeniopygia guttata), whether modifying an individual’s postnatal growth trajectory, using an ecologically-relevant scenario, results in consistent changes in adulthood mitochondrial functioning and if these relate to individual’s survival. Our results demonstrate that within individual age-related changes in mitochondrial function can be detected using RCBs in birds and they are evident by middle age. However, despite that some of the mitochondrial traits show individual consistency, we found no link between adult RBCs mitochondrial function and the pace of early growth or mid-term survival. Further research across the life course will help to interpret the implications of the observed changes for individual performance and its sensitivity to environmental changes.

Can growth conditions predict adulthood blood mitochondrial function and survival?

In nature, organisms are faced with constant nutritional options which fuel key life-history traits. Studies have shown that species can actively make nutritional decisions based on internal and external cues. Metabolism itself is underpinned by complex genomic interactions involving components from both nuclear and mitochondrial genomes. Products from these two genomes must coordinate how nutrients are extracted, used, and recycled. Given the complicated nature of metabolism, it is not well understood how nutritional choices are affected by mitonuclear interactions. This is under the rationale that changes in genomic interactions will affect metabolic flux and change physiological requirements. To this end we used a large Drosophila mitonuclear genetic panel, comprising 9 isogenic nuclear genomes coupled to 9 mitochondrial haplotypes, giving a total of 81 different mitonuclear genotypes. We use a capillary-based feeding assay to screen this panel for dietary preference between carbohydrate or protein. We find strong levels of sexual dimorphism for this trait, with females consuming a lot more protein than males. Furthermore, we find significant mitonuclear interactions modulating nutritional choices. This work gives us deeper insights on how key metabolic interactions can have large implications on behaviour.

Mitonuclear interactions modulate nutritional preference

Inspired by avians, BirdBot legged robot utilizes mechanical intelligence, physically embedded in its leg structure and control. BirdBot features mechanical joint coupling to coordinate joint movements and power. Networks of overlapping spring tendons act as joint flexors and extensors, and complex functions are achieved. BirdBot shows robust, agile, and energy-efficient locomotion with surprisingly little control effort.  In stance, a spring-tendon network is engaged and acts on all leg joints with joint extending torques. The orientation of the toe segment purely mechanically engages this spring-tendon network. In the swing phase, the leg is slack, with no or minimal joint torques acting. During the swing, a knee motor flexes the slack knee joint. The remaining, slack joints follow the motion coordinated by the robot's tendon network. Both modes are designed to provide the leg's stance and swing phase functions, reduce energy consumption, reduce control effort, and increase the robot leg's performance. In this work, we closely observe the function of the robot's distal toe-flexor spring-tendon, which spans the two most-distal joints. The toe-flexor spring-tendon continuously charges in the stance phase, accumulating energy. At the stance-swing transition, the tendon collapses the TMP joint. This action rapidly switches the leg from its loadable stance phase to its slack swing mode and also pushes the toe segment into a digital-flexed posture. Hence, besides its main function as a clutch trigger, the toe flexor-tendon also recuperates mechanical energy back in the leg. Energy stored in stance is released and supports rapidly flexing and leg protraction.  

Biarticular toe-flexor tendon and its function depending on locomotion speed

Biomechanical computational models have been comprehensively used to evaluate the physiology of locomotion within various extinct and extant vertebrate species. Such models offer enormous potential to explore parameters which would otherwise require invasive procedures to measure experimentally, however, it is currently unknown which functional inputs are integral to developing physiologically and biomechanically accurate models. Amongst the most important models inputs, though often ignored, are the force-velocity characteristics of the muscle actuators, which describe the relationship between force-generating capacity during shortening and lengthening. While the shortening velocity of muscles has been extensively studied, fibre behaviour during lengthening is less well understood, and as such are not included in many muscle driven simulations of vertebrate movement. Here, we aim to use the ‘fast’ extensor digitorum longus and ‘slow’ soleus from the mouse to explore if there exist phenotypic differences in the lengthening portion of the force-velocity relationship and explore the functional implications these parameters have on the outputs of a biomechanical simulation of mouse trotting. Using in-vitro isovelocity contractions that span ±5% of optimum fibre length on the force-length relationship, we show that several aspects of the force-velocity relationship differ significantly between the muscles. Applying these muscle-specific force-velocity curves to the musculoskeletal model, we show that the predicted lengthening and activation parameters were highly sensitive to these input parameters, as was muscle power. These data highlight the importance of accurately representing the force-velocity relationship of muscles for the generation of physiologically valid musculoskeletal models.



Functional consequences of the force-velocity relationship on simulated musculoskeletal dynamics

Limb movements and their control are widely studied using musculoskeletal models, but the results of biomechanical simulations can be heavily influenced by the choice of muscle model. In order to investigate how reaching movements are affected by intrinsic muscle properties, we have implemented a simple upper limb model with three joints, each controlled by an antagonistic muscle pair. By varying the characteristics of the muscles in the model, we have studied the consequences of using a simple activation dynamics model compared to a more realistic, higher-order activation model, and we have also investigated how activation dynamics interact with the muscles’ force-length-velocity characteristics to affect reaching outcomes. Our simulations suggest that accurate reaching movements can be achieved with a variety of muscle models, but compared to simple activation models, higher-order activation dynamics requires longer prediction from a forward model and gives rise to a higher level of unplanned co-contraction of antagonistic muscle pairs. In line with prior work, the force-length-velocity properties of muscles were observed to enable stabilisation and smoothing of limb movements in simulations as well as to promote accurate reaching performance with the high-order activation model. Overall, our simulation results support the use of simple muscle models when only the behavioural outcome is of interest. However, the choice of muscle model can become critical when addressing questions related to neuromuscular control, as models capable of capturing realistic muscle behaviour may also affect key features of the control problem.

The impact of muscle activation dynamics and other intrinsic properties on reaching movements: A simulation study

Elucidating transcriptional regulatory networks requires knowledge of transcription factor (TF) binding sites and their occupancy, ideally in a native chromatin context. Towards this goal, we have adapted the use of micrococcal nuclease (MNase) as a biochemical probe of fixed chromatin structure with emphasis on small DNA fragments from sub-nucleosome-sized particles. Specifically, we developed MNase-defined cistrome-Occupancy Analysis (MOA-seq) to achieve high-resolution genome-wide detection of TF-bound sites within accessible chromatin regions, illustrated with a proof of concept study on field-grown maize ear shoots (Savadel, Hartwig, et al., 2021; PLOS Genetics). Fragment center plotting focused the footprint detection down to small particles with an average peak size of ~30bp. These MOA footprints (MFs) exhibited (1) expression-correlated associations with gene promoters, (2) enrichment at known TF-binding sites, (3) enrichment at known long-range chromatin interaction sites, and (4) enrichment at known open chromatin regions defined by other non-MNase assays such as ATAC-seq. Motif analysis of 145,000 maize earshoot MFs led to the detection of 215 sequence motif families, which were merged into non-overlapping regions numbering just above 100,000 loci and covering just below 0.1% of the maize genome. We also used MOA-seq to examine genomic response to hypoxia in human cells. From these initial studies, we conclude that MOA-seq is ideal for genome-wide identification of cistrome occupancy at high resolution using a single antibody-free assay. The technology adds a valuable method to the genomic toolkit for pinpointing cis-regulatory candidates in agriculture and medicine.

MNase-based MOA-Seq Pinpoints Native Cistrome Occupancy at 30-bp Resolution Amid Accessible Chromatin

The interaction between chromatin and the nuclear lamina (NL) is intrinsically important to the establishment of three-dimensional chromatin architecture and spatiotemporal regulation of gene expression. However, critical regulators involved in this process are poorly understood in plants. Here, we report that Arabidopsis PNET2 and its two homologs are bona fide inner nuclear membrane proteins and integral components of the NL. PNET2s physically interact with the plant nucleoskeleton and engage nucleosome-enriched chromatin at the nuclear periphery. Loss of all three PNET2s leads to severely disrupted growth and development, concomitant activation of abiotic and biotic stress responses, and ultimate lethality in Arabidopsis. The pnet2 triple mutant also displays drastic transcriptome changes accompanied by a globally altered chromatin architecture revealed by HiC analysis. Our study identified PNET2 as an inner nuclear membrane (INM) component of the NL, which associates with chromatin and plays a critical role in orchestrating gene expression and chromatin organization in plants.











PNET2 is a component of the plant nuclear lamina and is required for proper genome organization and activity

Higher animals and plants are two evolutionary outcomes of complex multicellularity. An important driver
of cellular specialization and body plan expansion in both phyla was the evolution of vascular distribution
networks. Most prominently, the cardiovascular network of mammals distributes oxygen and nutrients
throughout the body, driven by the pumping action of the heart. By comparison, the vascular transport
systems of the dominant group of higher plants, the angiosperms, are fundamentally different. For instance,
transport of photosynthates in the phloem network is governed by dynamic source-sink relations and driven
by a pressure differential that builds up through locally controlled cellular osmolarity. The conducting
phloem channels are the so-called sieve tubes, which not only transport nutrients but also developmental
signals such as the phytohormone auxin. Sieve tubes are formed from interconnected individual sieve
elements that are meticulously aligned in cell files as they develop. Sieve elements represent a cellular
between-life-and-death state, because they lack numerous organelles, including the nucleus, and are
nurtured by their neighboring, so-called companion cells. The peculiar sieve element differentiation process
can be observed in growth apices. For example, in the root meristem of Arabidopsis thaliana seedlings their
development from stem cell into mature sieve element is laid out in a spatiotemporal gradient. My lab has
characterized a molecular network that guides this differentiation process through the interplay between
controlled transcellular auxin transport across developing sieve element cell files and autocrine receptor
kinase signaling pathways that respond to small peptide ligands. Key players in this network constitute a
molecular rheostat that finetunes transcellular auxin flux and maintains its pronounced cellular polarity
through a self-reinforcing mechanism. Our research suggests that this module is intimately connected to
the evolution of phloem, which enabled plants to effectively colonize land and thereby had a major impact
on the extant biosphere.

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