cAMP‐dependent reorganization of the Cajal bodies and splicing machinery in cultured Schwann cells

Abstract

It is well established that forskolin-induced elevation of cAMP results in activation of DNA synthesis in Schwann cell cultures. This promitotic response is partially mediated by the Cdk2, which is required for the transition from the G1 to the S phase of the cell cycle. In the present study, we analyze the effects of cAMP elevation in cultured Schwann cells on the transcriptional activity and on the organization of two nuclear compartments involved in pre-mRNA processing: Cajal bodies (CBs) and splicing factor compartments. Our immunofluorescence and quantitative studies show that forskolin treatment induces a 5.6-fold increase in the proportion of S phase Schwann cells, detected by a short pulse (20 min) of BrdU incorporation. This increase in DNA synthesis correlates with an activation of global transcription, as is indicated by the higher nuclear incorporation of BrU in nascent RNA. Forskolin treatment significantly increases the percentage of Schwann cells containing typical CBs, which concentrate spliceosomal snRNPs and the survival motor neuron (SMN) protein. This increase in the number of CBs closely correlates with the activation of transcription. Moreover, the occurrence of CBs is significantly higher in BrdU (+) cells than in BrdU (-) cells, indicating that entry in the S phase promotes the formation of CBs. During the S phase, Schwann cell nuclei display higher Cdk2 nuclear staining and concentrate this kinase in CBs. Forskolin also induces a redistribution of the pre-mRNA splicing factors in Schwann cells. Primary cultures of Schwann cells provide an excellent physiological model to demonstrate that the assembly of CBs is a transcription- and replication-dependent cellular event. Moreover, the S phase accumulation of Cdk2 observed in Schwann cells supports a functional link between CBs and DNA replication, which is mediated by the possible participation of CBs in the regulatory control of histone gene expression.

The Cajal body: a meeting place for spliceosomal snRNPs in the nuclear maze

“…Some of the proteins from this complex were localized in cleavage bodies, which are found in close proximity to CBs (Schul et al 1996). Moreover, Cajal and cleavage body appearance is cell cycle-dependent and cleavage bodies are found specifically adjacent to the Cajal bodies associated with histone genes (Carmo-Fonseca et al 1993;Fernandez et al 2002;Schul et al 1999b). Taken together, these observations support the proposal that CBs and/or adjacent cleavage bodies are sites of histone 3′-end processing.…”

Nucleolar targeting of coilin is regulated by its hypomethylation state

“…Canonical CBs are plurifunctional organelles involved in the biogenesis, transport, and recycling of snRNPs and snoRNPs (for a review, see Cioce and Lamond 2005;Stanek and Neugebauer 2005;Matera et al 2009). They are dynamic structures in many cell types, their number and size correlating with the phase of cell cycle and transcriptional activity (Lafarga et al 1991;Carmo-Fonseca et al 1993, Fernandez et al 2002Cioce and Lamond 2005). In fact, CBs represent dynamic steady-state systems that undergo rapid exchange of native components (Sleeman et al 1998;Misteli 2008).…”

Differentiating plant cells switched to proliferation remodel the functional organization of nuclear domains

“…The CBs are not the sites for splicing that occurs cotranscriptionally at transcription sites, but they represent the nuclear domain in which important maturing steps of splicing factors take place, before their recruitment for splicing sites (Spector, 1993;Proudfoot, 2000). The increase in CBs could reflect a cellular response to the high demand of mature splicing factors induced by the transcriptional activation (Ferná ndez et al, 2002) occurring in plant proliferating cells.…”

“…The snRNP immunofluorescence signal appeared more disperse in plant cells (Testillano et al, 1993). These splicing factors accumulate in Cajal bodies (Raska et al, 1990;Ferná ndez et al, 2002). Antibodies to splicing factors revealed these nuclear bodies as small bright spots (Moreno Díaz de la Espina et al, 1982;Testillano et al, 1993;Straatman and Schel, 2001).…”

“…3e). The increase in the number of CBs and the proportion of cells containing them has been closely correlated with transcriptional activation and cell cycle progression in various mammalian cells (Pena et al, 2001;Ferná ndez et al, 2002). The CBs are not the sites for splicing that occurs cotranscriptionally at transcription sites, but they represent the nuclear domain in which important maturing steps of splicing factors take place, before their recruitment for splicing sites (Spector, 1993;Proudfoot, 2000).…”