Immunohistochemical Detection of Ribosomal Transcription Factor UBF and AgNOR Staining Identify Apoptotic Events in Neoplastic Cells of Hodgkin's Disease and in Other Lymphoid Cells

Abstract

Ribosomal RNA synthesis is a key molecular process for understanding the mechanisms that drive cell proliferation. In this process, the upstream binding factor (UBF) is involved in regulating rDNA transcription at the nucleolus, together with RNA polymerase I. Recently, UBF was demonstrated to be a substrate for selective cleavage by specific proteases during apoptosis. Here we studied the expression of UBF in several cases of Hodgkin's disease (HD) by immunostaining and found it to be absent or clearly diminished in a high proportion of Reed-Sternberg cells and Hodgkin cells compared to small reactive lymphocytes. This result contrasted with labeling of those cells by the AgNOR technique, a marker of cell proliferation dependent on increased amounts of several proteins related to ribosome assembly. Disappearance of UBF and preservation of other NOR proteins is consistent with the pattern of selective proteolysis by caspases described in early stages of apoptosis. This correlates well with our results observed on induction of apoptosis in Jurkat cells treated with anti-FAS/APO-1 serum and with those in aged germinal center B-cells, in which UBF was no longer seen although the staining signal of other NOR proteins was maintained. These results support the concept that the rate of apoptosis is higher in neoplastic cells of HD than in the benign reactive lymphocyte population. Differential proteolysis of NOR proteins, as revealed by double staining of UBF and AgNOR, may prove valuable for identification of early stages of apoptosis in cytological and histopathological samples.

Different behavior of protein B23/nucleophosmin and UBF in HeLa cells during apoptosis

“…The findings of the present work are consistent with the data on camptothecin-induced of HeLa cells [5] and CD-95-induced apoptosis of Jurkat cells [4,8]. In these works the cleavage of UBF and the absence of B23 proteolysis were recorded, and total protein B23 content was unchanged even upon complete proteolysis of UBF [8].…”

“…In some works [4,5,8] UBF was shown to be an early target of the apoptosis-induced proteolysis, which depends on both cell types and apoptosis inducers. We decided to assess the behavior of UBF during apoptosis of HeLa cells induced by TNF-α and emetine mixture.…”

“…But what is known about apoptosis-dependent changes in the structure of UBF and nucleophosmin? The majority of available data indicate that in addition to translocation UBF undergoes apoptosis-dependent proteolysis [4,5,8]. Unfortunately in none of these studies the region of UBF proteolysis was characterized.…”

Nuclear localization of DEDD leads to caspase-6 activation through its death effector domain and inhibition of RNA polymerase I dependent transcription

“…34 It was demonstrated that UBF moves out of nucleoli during apoptosis whereas other nucleolar protein such as B23 stay within these structures. 35 This behavior has been interpreted as the result of the cleavage of UBF by caspases and has established UBF as a target that is inactivated early during apoptosis. Furthermore, it was recently shown that during apoptosis UBF localized to fibrogranular threaded bodies whereas fibrillarin, B23 and C23 remained present in micronuclei.…”

Changes in the status of nucleolus during long-term culturing of human HeLa cells

“…1, e), which attests to its probable degradation. By this sign A3 antigen corresponded to nucleolar Ag-NOR proteins, UBF (RNA polymerase I cofactor), and nucleolin (protein involved in the regulation of rDNA transcription and rRNA maturation) [9,11]. Hence, the appearance of cells with abnormal location of A3 antigen in HeLa culture was associated with increased sensitivity of the culture to suppression of total protein synthesis with anisomycin.…”