Macroautophagy allows for bulk degradation of cytosolic components in lysosomes. Overexpression of GFP/RFP-LC3/GABARAP is commonly used to monitor autophagosomes, a hallmark of autophagy, despite artifacts related to their overexpression. Here, we developed new sensors that detect endogenous LC3/GABARAP proteins at the autophagosome using an LC3-interacting region (LIR) and a short hydrophobic domain (HyD). Among HyD-LIR-GFP sensors harboring LIR motifs of 34 known LC3-binding proteins, HyD-LIR(TP)-GFP using the LIR motif from TP53INP2 allowed detection of all LC3/GABARAPs-positive autophagosomes. However, HyD-LIR(TP)-GFP preferentially localized to GABARAP/GABARAPL1-positive autophagosomes in a LIR-dependent manner. In contrast, HyD-LIR(Fy)-GFP using the LIR motif from FYCO1 specifically detected LC3A/B-positive autophagosomes. HyD-LIR(TP)-GFP and HyD-LIR(Fy)-GFP efficiently localized to autophagosomes in the presence of endogenous LC3/GABARAP levels and without affecting autophagic flux. Both sensors also efficiently localized to MitoTracker-positive damaged mitochondria upon mitophagy induction. HyD-LIR(TP)-GFP allowed live-imaging of dynamic autophagosomes upon autophagy induction. These novel autophagosome sensors can thus be widely used in autophagy research.

C-terminal fragments of Tar DNA-binding protein 43 (TDP-43) have been identified as the major pathological protein in several neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). However, how they affect cellular toxicity and neurodegeneration, including the modulation process remains unknown. This study revealed that the C-terminal fragment of TDP-43 (TDP-25) was localized primarily to mitochondria and caused abnormal mitochondrial morphology, inducing Parkin-mediated mitophagy. Also, we discovered that the knockdown of selective autophagy receptors, such as TAX1BP, Optineurin, or NDP52 caused TDP-25 accumulation, indicating that TDP-25 was degraded by mitophagy. Interestingly, myosin IIB, a nonmuscle type of myosin and actin-based motor protein, is mostly colocalized to TDP-25 associated with abnormal mitochondria. In addition, myosin IIB inhibition by siRNA or blebbistatin induced mitochondrial accumulation of insoluble TDP-25 and Tom20, and reduced neuronal cell viability. Our results suggest a novel role of myosin IIB in mitochondrial degradation of toxic TDP-25. Therefore, we proposed that regulating myosin IIB activity might be a potential therapeutic target for neurodegenerative diseases associated with TDP-43 pathology.

Arginine methylation plays crucial roles in many cellular functions including signal transduction, RNA transcription, and regulation of gene expression. Protein arginine methyl-transferase 8 (PRMT8), a unique brain-specific protein, is localized to the plasma membrane. However, the detailed molecular mechanisms underlying PRMT8 plasma membrane targeting remain unclear. Here, we demonstrate that the N-terminal 20 amino acids of PRMT8 are sufficient for plasma membrane localization and that oligomerization enhances membrane localization. The basic amino acids, combined with myristoylation within the N-terminal 20 amino acids of PRMT8, are critical for plasma membrane targeting. We also found that substituting Gly-2 with Ala [PRMT8(G2A)] or Cys-9 with Ser [PRMT8(C9S)] induces the formation of punctate structures in the cytosol or patch-like plasma membrane localization, respectively. Impairment of PRMT8 oligomerization/dimerization by C-terminal deletion induces PRMT8 mis-localization to the mitochondria, prevents the formation of punctate structures by PRMT8(G2A), and inhibits PRMT8(C9S) patch-like plasma membrane localization. Overall, these results suggest that oligomerization/dimerization plays several roles in inducing the efficient and specific plasma membrane localization of PRMT8.

The implementation of vehicle electrification and autonomous driving technologies has recently emphasized the importance of abrasion resistance and fuel efficiency of truck bus radial (TBR) tire treads that undergo high loads and long driving times. In this study, a functionalized liquid butadiene rubber (F-LqBR) was introduced to replace the treated distillate aromatic extracted (TDAE) oil as a way to improve abrasion resistance and fuel efficiency in the TBR tire tread compound and to solve the oil migration. First, radical polymerization was used to synthesize nonfunctionalized LqBR (N-LqBR) and amino-LqBR with an amine group at the chain ends. The synthesized LqBRs were then substituted in place of TDAE oil to manufacture carbon-black-filled natural rubber (NR) compounds and to evaluate their physical properties. The results show that LqBRs improved the migration resistance and enhanced the abrasion resistance by lowering the glass transition temperature (Tg) of the compound. In particular, amino-LqBR improved carbon black dispersion in the rubber matrix through a chemical bond between the functional group of the carbon black surface and the base rubber. In conclusion, amino-LqBR successfully served as a processing aid in a carbon black-filled NR compound while simultaneously enhancing its fuel efficiency and abrasion resistance.

As the world is shifting from internal combustion engine vehicles to electric vehicles in response to environmental pollution, the tire industry has been conducting research on tire performance to meet the requirements of electric vehicles. In this experiment, functionalized liquid butadiene rubber (F-LqBR) with triethoxysilyl groups at both ends was introduced into a silica-filled rubber compound as a substitute for treated distillate aromatic extract (TDAE) oil, and comparative evaluation was conducted according to the number of triethoxysilyl groups. The results showed that F-LqBRs improved silica dispersion in the rubber matrix through the formation of chemical bonds between silanol groups and the base rubber, and reduced rolling resistance by limiting chain end mobility and improving filler–rubber interaction. However, when the number of triethoxysilyl groups in F-LqBR was increased from two to four, self-condensation increased, the reactivity of the silanol groups decreased, and the improvement of properties was reduced. As a result, the optimized end functionality of triethoxysilyl groups for F-LqBR in silica-filled rubber compound was two. The 2-Azo-LqBR with the optimized functionality showed an improvement of 10% in rolling resistance, 16% in snow traction, and 17% in abrasion resistance when 10 phr of TDAE oil was substituted.

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