Immunoelectron microscopic study of polyamines in the gastrointestinal tract of rat

Abstract

Polyamines (PAs) are ubiquitous polycationic metabolites in the eukaryotic and prokaryotic cells and are believed to be intimately involved in the regulation of DNA, RNA, and protein biosynthesis. However, the subcellular localization of PAs has not yet been fully elucidated in a variety of cell types. In the present study, a pre-embedding indirect immunoperoxidase approach was used to define the fine structural localization of PAs in the gastrointestinal tract of rat, which was fixed with glutaraldehyde and the monoclonal antibody ASPM-29 specific for spermine (Spm) and spermidine (Spd). Examination by a transmission electron microscopy showed that the peroxidase end products were commonly and predominantly localized in the free and attached ribosomes of the rough endoplasmic reticulum (rER) in the active protein- or peptide-secreting cells, and in rapidly proliferating cells including the gastric chief cells, mucous neck cells, and intestinal crypt cells. The nuclei, mitochondria, and secretory vesicles were devoid of PAs. Of note is the new finding that PAs are also located even on the small number of ribosomes in the cytoplasm of the parietal cells and of the villus-tip cells, because these were the cell types that were found to be almost PA-negative at the light microscopic level. These results seem to be completely consistent with those recently obtained for rat neurons. Thus, the present study generalized the subcellular localization of PAs on the ribosomes, and demonstrated that PAs are one of the components of biologically active ribosomes, possibly in any type of cell, that are closely involved in the translation processes of protein biosynthesis.

Polyamines in spermatocytes and residual bodies of rat testis

“…In thusdeWned residual bodies, RNA colocalized together with PAs and ultrastructural immunocytochemistry demonstrated that such colocalization reXected an association between PAs and ribosomes. This is in good agreement with our previous results showing PA localization on ribosomes in neurons and gastrointestinal epithelial cells (Fujiwara et al 1998;Tanabe et al 2004;Shin et al 2006). Since residual bodies are formed from maturing spermatids it is a triXe surprising that the latter cells stained weaklier for PAs.…”

“…Immunocytochemical localization of PAs may help to unravel their true biological roles by revealing their exact cellular and subcellular localization. We have previously demonstrated that PAs are localized in association with ribosomes in neurons and gastrointestinal epithelial cells (Fujiwara and Masuyama 1995;Fujiwara et al 1998;Tanabe et al 2004;Shin et al 2006): localization in agreement with biochemical studies suggesting a role for PAs in protein synthesis Tabor 1984, 1985;Pegg 1986;Yoshida et al 2004).…”

“…The monoclonal antibody ASPM-29 (IgG1 sub-isotype) used in this study was from the same batch as used in previous reports (Fujiwara and Masuyama 1995;Fujiwara et al 1998;Shin et al 2006). The speciWcity of the immunocytochemical method that utilized the ASPM-29 monoclonal antibody has been characterized extensively (Fujiwara and Masuyama 1995;Fujiwara et al 1996Fujiwara et al , 1997).…”

Immunocytochemical demonstration of polyamines in nucleoli and nuclei

“…Using the same principle, we have previously demonstrated that PAs occur associated with both free and ER-bound ribosomes in the cytoplasm of any cell type (Fujiwara and Masuyama 1995;Fujiwara et al 1998a;Tanabe et al 2004;Shin et al 2006Shin et al , 2007. Hougaard et al (1987) have reported on nuclear localization of PAs in chick erythrocytes, and more recently, Johnson et al (2004) reported on nuclear staining for PAs in cell lines, although their data were not supported by speciWcity (absorption) controls.…”

“…This ICC system has proved useful for detecting PAs in cells and tissues (Fujiwara et al 1996(Fujiwara et al , 1997a. Moreover, by immunoelectron microscopy, this ICC system revealed that PAs are associated with both free and ER-bound ribosomes in all cell types investigated (Fujiwara et al 1998a;Tanabe et al 2004;Shin et al 2006Shin et al , 2007. Since ribosomes originate from nucleoli, we asked ourselves whether PAs also were associated with nucleoli and other regions of the nucleus.…”