Identifying mRNAs Residing in Myelinating Oligodendrocyte Processes as a Basis for Understanding Internode Autonomy

Abstract

In elaborating and maintaining myelin sheaths on multiple axons/segments, oligodendrocytes distribute translation of some proteins, including myelin basic protein (MBP), to sites of myelin sheath assembly, or MSAS. As mRNAs located at these sites are selectively trapped in myelin vesicles during tissue homogenization, we performed a screen to identify some of these mRNAs. To confirm locations, we used real-time quantitative polymerase chain reaction (RT-qPCR), to measure mRNA levels in myelin (M) and ‘non-myelin’ pellet (P) fractions, and found that five (LPAR1, TRP53INP2, TRAK2, TPPP, and SH3GL3) of thirteen mRNAs were highly enriched in myelin (M/P), suggesting residences in MSAS. Because expression by other cell-types will increase p-values, some MSAS mRNAs might be missed. To identify non-oligodendrocyte expression, we turned to several on-line resources. Although neurons express TRP53INP2, TRAK2 and TPPP mRNAs, these expressions did not invalidate recognitions as MSAS mRNAs. However, neuronal expression likely prevented recognition of KIF1A and MAPK8IP1 mRNAs as MSAS residents and ependymal cell expression likely prevented APOD mRNA assignment to MSAS. Complementary in situ hybridization (ISH) is recommended to confirm residences of mRNAs in MSAS. As both proteins and lipids are synthesized in MSAS, understanding myelination should not only include efforts to identify proteins synthesized in MSAS, but also the lipids.

Recognizing the Beauty of Myelination through Identifying Sites of Phospholipid, Protein, and RNA Syntheses and Characterizing Their Movements in Myelin Internodes

“…These findings provide morphological visualization of distinct locations where syntheses of these proteins occur. Furthermore, in addition MBP, myelinating oligodendrocytes, and likely Schwann cells, contain other mRNAs that co-localize with MBP mRNA in processes, indicating that silver grains in SCC/CB, not only represent MBP synthesis, but other proteins important for myelination [10][11][12]. Taken together, these studies demonstrate spatial separations of sites where different myelin-destined proteins are synthesized, information not extracted in readouts typically used to assess myelination.…”

“…Autoradiographic studies could readily be conducted, as was done for choline lipid metabolism in frog sciatic nerves [20], using species of birds [64], reptiles [65,66], amphibians (Xenopus [67,68] and Rana [20,69]), teleost (trout [70]) and cartilaginous (little skate [71]) fishes used previously in studies of peripheral nerve myelination. Our endeavors in identifying mRNAs that co-localize with MBP mRNA in oligodendrocyte processes [10,12] came in part from finding that unlike in other species, in elasmobranchs, MBP mRNA is not enriched in oligodendrocyte processes [72].…”

Perspective Strategies for Interventions in Parkinsonism: Remedying the Neglected Role of TPPP

“…Recently, single-nucleus RNA sequencing revealed a high level of SNCA transcripts in inhibitory neurons and OLG progenitor cells but only a low level of these transcripts in excitatory neurons and mature OLGs [40]. In the case of TPPP, its mRNA is present in neurons but the protein itself is only present in OLGs [17,41]. Hence, possible pathological factors include a failure to inhibit translation of the inappropriate mRNA (an inhibition that should occur in physiological conditions) and/or a failure to degrade the newly synthesized protein; these possibilities would be consistent with the absence of the SYN protein in mature OLGs despite the presence of its mRNA.…”