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. 2019 Apr 1;12(7):1064.
doi: 10.3390/ma12071064.

Fluorescence-Based Analysis of Noncanonical Functions of Aminoacyl-tRNA Synthetase-Interacting Multifunctional Proteins (AIMPs) in Peripheral Nerves

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Free PMC article

Fluorescence-Based Analysis of Noncanonical Functions of Aminoacyl-tRNA Synthetase-Interacting Multifunctional Proteins (AIMPs) in Peripheral Nerves

Muwoong Kim et al. Materials (Basel). .
Free PMC article

Abstract

Aminoacyl-tRNA synthetase-interacting multifunctional proteins (AIMPs) are auxiliary factors involved in protein synthesis related to aminoacyl-tRNA synthetases (ARSs). AIMPs, which are well known as nonenzymatic factors, include AIMP1/p43, AIMP2/p38, and AIMP3/p18. The canonical functions of AIMPs include not only protein synthesis via multisynthetase complexes but also maintenance of the structural stability of these complexes. Several recent studies have demonstrated nontypical (noncanonical) functions of AIMPs, such as roles in apoptosis, inflammatory processes, DNA repair, and so on. However, these noncanonical functions of AIMPs have not been studied in peripheral nerves related to motor and sensory functions. Peripheral nerves include two types of structures: peripheral axons and Schwann cells. The myelin sheath formed by Schwann cells produces saltatory conduction, and these rapid electrical signals control motor and sensory functioning in the service of survival in mammals. Schwann cells play roles not only in myelin sheath formation but also as modulators of nerve degeneration and regeneration. Therefore, it is important to identify the main functions of Schwann cells in peripheral nerves. Here, using immunofluorescence technique, we demonstrated that AIMPs are essential morphological indicators of peripheral nerve degeneration, and their actions are limited to peripheral nerves and not the dorsal root ganglion and the ventral horn of the spinal cord.

Keywords: Schwann cells; Wallerian degeneration; aminoacyl-tRNA synthetase-interacting multifunctional proteins (AIMPs); fluorescence-based analysis; noncanonical functions.

Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
mRNA expression patterns of aminoacyl-tRNA synthetase-interacting multifunctional proteins (AIMPs) under normal and abnormal conditions of peripheral nerves. (a) Real-time quantitative PCR (qPCR) results showed mRNA expressions of AIMPs in sciatic nerves. Relative mRNA expressions of AIMPs were calculated by fold change; 1 was expression level of control sample (Con, not injured nerves). Fold changes of postinjury samples (3 days, 1 week, 2 weeks, 4 weeks) were derived by mRNA expression level of each sample divided by that of Con (n = 3, * p < 0.05, ** p < 0.01, *** p < 0.001, ns; p > 0.05). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as loading control. (b) Semi-PCR results showed mRNA expressions of AIMPs in sciatic nerve.
Figure 2
Figure 2
Protein expression patterns of AIMPs under normal and abnormal conditions of peripheral nerves. (a) Western blot analysis showed protein expression patterns of AIMPs in sciatic nerves. (b) Relative intensities of protein expressions of AIMPs were calculated by fold change; 1 was expression level of Con. Fold changes of postinjury samples (3 days, 1 week, 2 weeks, 4 weeks) were derived by protein expression level of each sample divided by that of Con (n = 3, * p < 0.05, ** p < 0.01, *** p < 0.001, ns; p > 0.05). β-actin was used as loading control.
Figure 3
Figure 3
Morphological expression patterns of AIMPs under normal and abnormal conditions of peripheral nerves. (a) Immunostaining showed colocalization of AIMPs (red) with S100 (green, Schwann cell marker) in teased sciatic nerves. Expressions of AIMPs were maximized at 1 week and gradually decreased from 1 week after injury. Scale bar = 100 μm. (b) Relative intensities of AIMPs were calculated by strength of each AIMP signal; relative intensity of 1 week was 100. Relative intensities of postinjury samples (Con, 3 days, 2 weeks, 4 weeks) were derived by strength of signal of each sample divided by that of 1 week (n = 3, * p < 0.05, ** p < 0.01, *** p < 0.001, ns; p > 0.05).
Figure 4
Figure 4
AIMPs are increased in Schwann cells under dysfunctional condition of peripheral nerves. (ac) Immunostaining showed colocalization of AIMP1-3 (red) with S100 (green), not neurofilament (NF; green, axonal marker) after injury. Scale bar = 50 μm. (d,e) Cell counts were derived by S100/AIMPs double-positive cells or NF/AIMPs double-positive cells at Con or 3 days postinjury (n = 3, * p < 0.05, ** p < 0.01, *** p < 0.001, ns; p > 0.05).
Figure 5
Figure 5
Alteration of AIMPs expression is limited to Schwann cells under dysfunctional condition of peripheral nerves. (a) Immunostaining showed colocalization of AIMPs (red) with NeuN (green, neuronal marker) in dorsal root ganglia (DRGs). Scale bar = 100 μm. (b) Immunostaining showed colocalization of AIMPs (red) with S100 (green, satellite cell marker) in DRG. Scale bar = 50 μm. (c) Western blot analysis showed expression patterns of AIMPs in DRGs with or without nerve injury. β-actin was used as loading control. (d) Relative intensities of immunostaining of AIMPs were calculated by fold change of the injured DRG compared to that of Con; Con was 100 (n = 3, ns; p > 0.05).
Figure 6
Figure 6
Alteration of AIMPs expressions is similar to p75 neurotrophin receptor (NTR) as a marker of Schwann cell trans-dedifferentiation. (a) Immunostaining showed colocalization of AIMPs (red) with p75NTR (green) in sciatic nerve fibers. Scale bar = 100 μm. (b) Relative intensities of immunostaining of AIMPs and p75NTR were calculated by fold change of the degenerating and regenerating nerve fibers compared to that of samples at 1 wk after nerve injury, time-course-dependently (n = 3, p < 0.001).

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