PMP22 mRNA Research Articles

Differential Evolution and Expression of Murine Peroxisomal Membrane Protein Genes

Gene segments encoding the 70 and 22-kDa peroxisomal membrane proteins (PMP) have been characterized in mice and compared with other peroxisomal proteins in terms of evolution and expression. The mouse PMP22 gene sequence predicts A 16G and I 136V substitutions that agree with those defined by cyanogen bromide cleavage analysis, providing additional evidence that this gene encodes the major 22-kDa membrane protein visualized by SDS-polyacrylamide electrophoresis. Mammalian PMP22 genes exhibit higher evolutionary rates (0.17% amino acid substitution per million years) than PMP35 (0.14%), PMP70 (0.07%), or catalase genes (0.13%). PMP70 gene regions are conserved throughout vertebrate phylas based on Southern analysis, while PMP22 sequences were only detected in rodents. Amino acid substitutions are clustered in both PMP22 and PMP70 genes, and their pattern supports membrane topologies derived from hydropathy profiles. Northern blot analysis identifies single mRNAs of 4.6 kb (PMP70), 1.4 kb (PMP22), and 2.3 kb (catalase) in several rodent tissues. Quantitative or competitive RT-PCR assays detected two- to three-fold greater numbers of catalase mRNA molecules relative to PMP mRNA molecules in brain, liver, and kidney; PMP22 and PMP35 mRNAs were two-fold more abundant than PMP70 mRNA in these tissues. Steady-state levels of PMP22 mRNA were highest in rodent liver kidney, spinal cord, and duodenum with low levels in colon, adrenal, thymus, and spleen. We conclude that PMP genes exhibit independent evolutionary rates and tissue regulation suggesting that they have unique roles in peroxisome biogenesis aod tissue differentiation.

Peripheral Myelin Protein‐22 is Expressed in Rat and Mouse Brain and Spinal Cord Motoneurons

Peripheral myelin protein PMP-22 is expressed by Schwann cells and is a constituent of peripheral nervous system (PNS) myelin. Two PMP-22 transcripts, SR13 and CD25, differing in their 5' non-coding sequences have been described. SR13 is present both in the PNS and in non-neural tissue, whereas CD25 mRNA is almost exclusively expressed in Schwann cells. PMP-22 mRNA is also present in the central nervous system (CNS), but at much lower levels than in the PNS. We have investigated the regional distribution of PMP-22 mRNA in the rat and mouse CNS by the reverse transcriptase-polymerase chain reaction method, using oligonucleotide primers specific for the SR13 or CD25 transcripts. SR13 mRNA was detected in all the CNS regions analysed, whereas the CD25 message was present only in the brainstem and the spinal cord. Localization of the PMP-22 transcripts, determined by in situ hybridization in 21 day-old animals, showed selective expression in the motor nuclei. The PMP-22 signal was very weak in the nuclei of the oculomotor and trochlear nerves and absent in the nucleus of the abducens nerve. A strong PMP-22 signal was observed in the motor nuclei of the trigeminal, facial, ambigus, vagus, hypoglossal and accessory spinal nerves and in the ventral horn of the spinal cord. The PMP-22-positive cells were identified as motoneurons on the basis of topographic and morphological criteria, as well as immunolabelling with neuron-specific antibodies.(ABSTRACT TRUNCATED AT 250 WORDS)

Retroviral-mediated gene transfer of the peripheral myelin protein PMP22 in Schwann cells: modulation of cell growth.

The peripheral myelin gene PMP22 is the rat and human homologue of the murine growth arrest-specific gene gas3. Besides a putative role of PMP22 in myelination, a regulatory function in cell growth has been suspected. Here we have investigated both the expression of PMP22 during cell cycle progression of cultured rat Schwann cells and the influence of altered levels of PMP22 on Schwann cell growth. When resting cells were stimulated to begin the cell cycle, the regulation of PMP22 mRNA resembled the growth arrest-specific pattern of gas3 expression observed previously in NIH3T3 fibroblasts. To prove a growth regulatory function of PMP22, we generated Schwann cell cultures by infection with retroviral PMP22 expression vectors that constitutively expressed PMP22 cDNA sequences, in either the sense or antisense orientation. Transduced cells carrying the sense construct overexpressed PMP22 mRNA and protein, whereas in cells infected with an antisense PMP22 expression vector PMP22 mRNA levels were reduced markedly. Altered levels of PMP22 significantly modulated Schwann cell proliferation, as judged by 5-bromo-2'-deoxy-uridine incorporation into replicated DNA. In asynchronously dividing cultures enhanced expression of PMP22 decreased DNA synthesis to 60% of the control level. Conversely, reduced levels of PMP22 mRNA led to enhanced DNA synthesis of approximately 150%. Further cell cycle analyses by flow cytometry revealed that overexpression of PMP22 delayed serum- and forskolin-stimulated entry of resting Schwann cells from G0/G1 into the S + G2/M phases by approximately 8 h, whereas underexpression of PMP22 mRNA slightly increased the proportion of cells that entered the S + G2/M phases.(ABSTRACT TRUNCATED AT 250 WORDS)

Coexpression of PMP22 gene with MBP and P0 during de novo myelination and nerve repair.

The recently cloned PMP22 gene, the rat variant of the murine growth arrest-specific gene gas3, encodes a new 22 kD integral membrane glycoprotein of peripheral myelin. By means of in situ hybridization and immunohistochemistry, we have (1) analyzed PMP22 expression in myelinated and nonmyelinated peripheral nerves, and (2) compared the spatio-temporal changes in the expression of PMP22 mRNA with the expression of the myelin genes P0 and MBP (myelin basic protein) in developing as well as degenerating and regenerating sciatic nerve of rat. (3) We further investigated the expression of PMP22 mRNA by Northern blot in cultured Schwann cells maintained under different conditions of cell growth and arrest. Expression of PMP22 mRNA is restricted to Schwann cells of myelinated peripheral nerve. Transection of sciatic nerve in adult rat leads to a simultaneous and rapid decline in both PMP22 and P0 mRNA to nondetectable levels in the degenerating distal stump. When a demyelinated and axon-free distal stump, as indicated by the lack of MBP and neurofilament immunoreactivity, was reanastomosed to its proximal counterpart, the coordinated reexpression of PMP22 and MBP succeeded axonal regeneration through the distal segment with a delay of 1-2 weeks. As in regenerating nerve, a striking synchrony of expression of PMP22 and P0 transcripts, as well as MBP immunoreactivity, could be observed during sciatic nerve development. Further, in vitro evidence suggests that, unlike NIH3T3-fibroblasts, expression of PMP22/gas3 is not strictly growth arrest-specific in Schwann cells.(ABSTRACT TRUNCATED AT 250 WORDS)