Tm7

TGN46 + TM7

Fig. 8. Transmembrane domain 3 of MNK contains a trans-Golgi retention signal. Colocalization studies comparing the trans-Golgi resident protein TGN46 with CD8 fusion proteins containing transmembrane domain 3 or 7 were performed. Wild-type CD8 is localized to the plasma membrane (B) and does not colocalize with TGN46 (A,C). The staining pattern of the fusion protein of CD8 containing transmembrane domain 3 of MNK (E) is almost identical to that of TGN46 (D). The degree of overlap is confirmed by the bright signal (F). The staining pattern of the fusion protein of CD8 containing transmembrane domain 7 of MNK is characteristic of the endoplasmic reticulum (H) and does not overlap with TGN46 (I). (Reprinted from ref. 16 with permission from Oxford University Press.)

Fig. 8. Transmembrane domain 3 of MNK contains a trans-Golgi retention signal. Colocalization studies comparing the trans-Golgi resident protein TGN46 with CD8 fusion proteins containing transmembrane domain 3 or 7 were performed. Wild-type CD8 is localized to the plasma membrane (B) and does not colocalize with TGN46 (A,C). The staining pattern of the fusion protein of CD8 containing transmembrane domain 3 of MNK (E) is almost identical to that of TGN46 (D). The degree of overlap is confirmed by the bright signal (F). The staining pattern of the fusion protein of CD8 containing transmembrane domain 7 of MNK is characteristic of the endoplasmic reticulum (H) and does not overlap with TGN46 (I). (Reprinted from ref. 16 with permission from Oxford University Press.)

that there is a signal within a 38 amino acid (aa) stretch containing the TM3 of MNK sufficient to retain a protein at the TGN.

Interestingly, an alternatively spliced MNK product that skips exon 10 (containing TM3 and TM4) is present in low amounts in normal individuals. The protein from this transcript was localized to the endoplasmic reticulum (30) and the authors suggest that this arises because this aberrant splicing prevents correct protein folding or interferes with a Golgi "localization domain" (30).

In summary, we identified a signal sufficient for the retention of fusion proteins at the TGN in a 38 amino acid stretch containing TM3, inferring that TM3 is an essential element in the localization of MNK to the TGN (16). It is possible that the retentive qualities of TM3 are compromised on addition of copper, resulting in an increased proportion of the protein moving to the cell surface.

Fig. 9. C-terminus deletions of MNK. MCF1 (Menkes cytoplasmic fragment 1) contains the terminal 92 amino acids of MNK. This, along with the deleted fragment (MCF2), contains four putative internalisation motifs; three of the dileucine type, LL1, LL3, and LL4 and one of the tyrosine type, Y2 (YSRA). LL4 is the terminal 24-amino-acid fragment containing only 1 internalization type motif LL4 (31).

Fig. 9. C-terminus deletions of MNK. MCF1 (Menkes cytoplasmic fragment 1) contains the terminal 92 amino acids of MNK. This, along with the deleted fragment (MCF2), contains four putative internalisation motifs; three of the dileucine type, LL1, LL3, and LL4 and one of the tyrosine type, Y2 (YSRA). LL4 is the terminal 24-amino-acid fragment containing only 1 internalization type motif LL4 (31).

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