Biology subjects

Willingham, T. B.

Publications and source records attributed to Willingham, T. B..

5 recordsLinked to original sources

bioRxiv2024

Impact of Capillary and Sarcolemmal Proximity on Mitochondrial Structure and Energetic Function in Skeletal Muscle

Mitochondria within skeletal muscle cells are considered to be located either between the muscle contractile apparatus (interfibrillar mitochondria, IFM) or in large pools beneath the cell membrane (subsarcolemmal mitochondria, SSM), with several structural and functional differences reported between IFM and SSM. However, recent 3D imaging studies suggest that proximity to capillaries embedded in sarcolemmal grooves, rather than proximity to the sarcolemma itself, may drive the accumulation of mitochondria near the cell periphery (paravascular mitochondria, PVM). To evaluate the impact of capillary versus sarcolemmal proximity, we compared the structure and function of skeletal muscle mitochondria located either in large pools lateral to embedded capillaries (PVM), adjacent to the sarcolemma but not in PVM pools (SSM), or interspersed between sarcomeres (IFM). Mitochondrial morphology and interactions were assessed by 3D electron microscopy coupled with machine learning segmentation while mitochondrial energy conversion was assessed by two-photon microscopy of mitochondrial membrane potential, content, calcium, NADH redox and flux in live, intact cells. Structurally, while PVM and SSM were similarly larger than IFM, PVM were more compact and had greater mitochondrial connectivity compared to both IFM and SSM. Functionally, PVM had similar or greater basal NADH flux compared to SSM and IFM, respectively, despite a more oxidized NADH pool and a greater membrane potential, signifying a greater activation of the electron transport chain in PVM. Together, these data indicate proximity to capillaries has a greater impact on mitochondrial energy conversion and distribution in skeletal muscle than the sarcolemma alone.

physiology↗

bioRxiv2022

Ablation of Tmem65 Causes Lethal Mitochondrial Encephalomyopathy in Mouse

Transmembrane protein 65 (TMEM65) depletion in a patient carrying a homozygous variant in the Tmem65 splice site resulted in severe mitochondrial encephalomyopathy, indicating the clinical importance of TMEM65. However, the function of TMEM65 remains unknown. Here, we generated a TMEM65 reporter mouse as well as whole-body and tissue-specific Tmem65 knockout (KO) mice to investigate the localization and function of TMEM65. We show that TMEM65 is localized to mitochondria in heart, skeletal muscle, and throughout the brain. Both whole-body and nervous system-specific Tmem65 KO result in severe growth retardation and sudden death following seizures ~3 weeks after birth, indicating TMEM65 is indispensable for normal brain function. In addition, we find that skeletal muscle-specific Tmem65 KO leads to progressive, adult-onset myopathy preceded by elevated mitochondrial calcium levels despite unaltered expression of known mitochondrial or cellular calcium handling proteins. Consistently, we demonstrate that ablation of TMEM65 results in a loss of sodium-dependent mitochondrial calcium export. Finally, we show that blocking mitochondrial calcium entry through removal of the mitochondrial calcium uniporter (MCU) rescues the early lethality of whole-body TMEM65 ablation. Our data not only reveal the essential role of TMEM65 in mammalian physiology, but also suggest modulating mitochondrial calcium may offer a potential therapeutical approach to address defects associated with TMEM65 misexpression.

physiology↗

bioRxiv2022

Mitochondrial Network Configuration Influences Sarcomere and Myosin Filament Structure in Striated Muscles

Sustained muscle contraction occurs through interactions between actin and myosin filaments within sarcomeres and requires a constant supply of adenosine triphosphate (ATP) from nearby mitochondria. However, it remains unclear how different physical configurations between sarcomeres and mitochondria alter the energetic support for contractile function. Here, we show that sarcomere cross-sectional area (CSA) varies along its length in a cell type-dependent manner where the reduction in Z-disk CSA relative to the sarcomere center is closely coordinated with mitochondrial network configuration in flies, mice, and humans. Further, we find myosin filaments near the sarcomere periphery are curved relative to interior filaments with greater curvature for filaments near mitochondria compared to sarcoplasmic reticulum. Finally, we demonstrate smaller myosin filament lattice spacing at filament ends than filament centers in a cell type-dependent manner. These data suggest both sarcomere structure and myofilament interactions are influenced by the location and orientation of mitochondria within muscle cells.

cell biology↗

bioRxiv2021

Regulation of the Evolutionarily Conserved Muscle Myofibrillar Matrix by Cell Type Dependent and Independent Mechanisms

Skeletal muscles play a central role in human movement through forces transmitted by contraction of the sarcomere. We recently showed that mammalian sarcomeres are connected through frequent branches forming a singular, mesh-like myofibrillar matrix. However, the extent to which myofibrillar connectivity is evolutionarily conserved as well as mechanisms which regulate the specific architecture of sarcomere branching remain unclear. Here, we demonstrate the presence of a myofibrillar matrix in the tubular, but not indirect flight (IF) muscles within Drosophila melanogaster. Moreover, we find that loss of transcription factor H15 increases sarcomere branching frequency in the tubular jump muscles, and we show that sarcomere branching can be turned on in IF muscles by salm-mediated conversion to tubular muscles. Finally, we demonstrate that neurochondrin knockdown results in myofibrillar connectivity in IF muscles without conversion to tubular muscles. These data indicate an evolutionarily conserved myofibrillar matrix regulated by both cell-type dependent and independent mechanisms.

cell biology↗

bioRxiv2019

MUSCLE MITOCHONDRIAL CAPACITY AND ENDURANCE IN ADULTS WITH TYPE 1 DIABETES

The impact of type 1 diabetes (T1D) on muscle endurance and oxidative capacity is currently unknown. PurposeMeasure muscle endurance and oxidative capacity of adults with T1D compared to controls. MethodsA cross-sectional study design with a control group was used. Subjects (19-37 years old) with T1D (n=17) and controls (n=17) were assessed with hemoglobin A1c (HbA1c) and casual glucose. Muscle endurance was measured with an accelerometer at stimulation frequencies of 2, 4, and 6 Hz for a total of nine minutes. Mitochondrial capacity was measured using near-infrared spectroscopy after exercise as the rate constant of the rate of recovery of oxygen consumption. ResultsT1D and control groups were similar in age, sex, height, and race. The T1D group had slightly higher BMI values and adipose tissue thickness over the forearm muscles. Casual glucose was 150{+/-}70 mg/dL for T1D and 98{+/-}16 mg/dL for controls (P=0.006). HbA1c of T1D subjects was 7.1{+/-}0.9% and 5.0{+/-}0.4% for controls (P<0.01). Endurance indexes at 2, 4, and 6 Hz were 94.5{+/-}5.2%, 81.8{+/-}8.4%, and 68.6{+/-}13.5% for T1D and 94.6{+/-}4.1%, 85.9{+/-}6.3%, and 68.7{+/-}15.4% for controls (p = 0.97, 0.12, 0.99, respectively). There were no differences between groups in mitochondrial capacity (T1D= 1.9{+/-}0.5 min-1 and control=1.8{+/-}0.4 min-1, P=0.29) or reperfusion rate (T1D= 8.8{+/-}2.8s and control=10.3{+/-}3.0s, P=0.88). There were no significant correlations between HbA1c and either muscle endurance, mitochondrial capacity or reperfusion rate. ConclusionsAdults with T1D did not have reduced oxidative capacity, muscle endurance or muscle reperfusion rates compared to controls. HbA1c also did not correlate with muscle endurance, mitochondrial capacity or reperfusion rates. Future studies should extend these measurements to older people or people with poorly-controlled T1D.

physiology↗