Many genes have been described and characterized which result in alternative polyadenylation site use at the 3'-end of their mRNAs based on the cellular environment. In this survey and summary article 95 genes are discussed in which alternative polyadenylation is a consequence of tandem arrays of poly(A) signals within a single 3'-untranslated region. An additional 31 genes are described in which polyadenylation at a promoter-proximal site competes with a splicing reaction to influence expression of multiple mRNAs. Some have a composite internal/terminal exon which can be differentially processed. Others contain alternative 3'-terminal exons, the first of which can be skipped in some cells. In some cases the mRNAs formed from these three classes of genes are differentially processed from the primary transcript during the cell cycle or in a tissue-specific or developmentally specific pattern. Immunoglobulin heavy chain genes have composite exons; regulated production of two different Ig mRNAs has been shown to involve B cell stage-specific changes in trans -acting factors involved in formation of the active polyadenylation complex. Changes in the activity of some of these same factors occur during viral infection and take-over of the cellular machinery, suggesting the potential applicability of at least some aspects of the Ig model. The differential expression of a number of genes that undergo alternative poly(A) site choice or polyadenylation/splicing competition could be regulated at the level of amounts and activities of either generic or tissue-specific polyadenylation factors and/or splicing factors.

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Immunoglobulin (Ig) heavy-chain proteins exist in two forms, differing only at their carboxyl termini. The membranebound antigen receptor is found on the surface of early or memory B cells, while the secreted form is produced by plasma cells. The mRNAs for the two protein forms are derived from a single pre-mRNA that is alternatively processed at the 3Ј end (1,8,44,45). The 3Ј end of the secretion-specific (sec) mRNA is encoded by the last constant-region exon; the sec poly(A) site is about 100 nucleotides downstream of the last constantregion exon. The 3Ј end of the membrane-specific (mb) mRNA is encoded by a large portion of the last constant-region exon and by two downstream exons, M1 and M2. mb mRNA is produced when splicing of the last constant-region exon to M1 takes place using a 5Ј splice site within the last constant-region exon and polyadenylation occurs at the poly(A) site at the end of M2.The ratio of sec to mb mRNA is indicative of a B cell's developmental stage (reviewed in reference 18). Early B, pre-B, and memory cells and their tumor analogs, lymphomas, make approximately equal amounts of sec and mb mRNA. Plasma cells and their tumor counterparts, myelomas, make 10-to 100-fold more sec mRNA than mb mRNA. While translational and posttranslational control mechanisms play a small role in contributing to the sec-versus-mb protein ratios of Igproducing cells, the major contribution to the phenotype comes from differential production of sec and mb mRNAs (13,24,25,41). Studies of the mouse Ig ␥2b locus have shown that transcription termination occurs downstream of the mb poly(A) site in both early and late-stage B cells (11). This finding is in contrast to some studies of the Ig transcription unit (13,19), in which differential transcription termination has been suggested as a possible mechanism for the switch in poly(A) site use during B-cell development. The stability of both ␥ sec and ␥ mb mRNAs increases in late-stage or plasma cells, and differential mRNA stability therefore cannot account for the major shift in the ratio of the two mRNAs (33).The key point in understanding posttranscriptional processing of Ig heavy-chain mRNAs is that splicing of the last constant-region exon to M1 and polyadenylation at the sec poly(A) site are two mutually exclusive events. The balance between the two is reflected in the final sec-to-mb mRNA ratio and could potentially be controlled by regulation of either event. However, when the efficiency of splicing of the last constant-region exon to M1 exons was examined in either the or ␥ gene, there was no change in early versus late-stage B cells or in lymphoid versus nonlymphoid cell lines (3,38,40,56), indicating that splicing is a constitutive, nonregulated step. In contrast, experiments using stable transfections have shown that regulated expression of the mouse Ig ␥2b gene is influenced by poly(A) site order and strength (26).The biochemistry of cleavage and polyadenylation and the sequences required in cis in the pre-mRNA for 3Ј-end formation have been well chara...

The amount of the 64-kDa subunit of polyadenylation͞cleavage stimulatory factor (CstF-64) increases 5-fold during the G 0 to S phase transition and concomitant proliferation induced by serum in 3T6 fibroblasts. Higher levels of CstF-64 result in an increase in CstF trimer. The rise in CstF-64 occurs at a time when the amount of poly(A)-containing RNA rose at least 5-8 fold in the cytoplasm. Primary human splenic B cells, resting in G 0 , show a similar 5-fold increase in CstF-64 when cultured under conditions inducing proliferation (CD40 ligand exposure). Therefore, the increase in CstF-64 is associated with the G 0 to S phase transition. As B cell development progresses, RNA processing changes occur at the Ig heavy chain locus resulting in a switch from the membrane-to the upstream secretory-specific poly(A) site. Treating resting B cells with agents triggering this switch in Ig mRNA production along with proliferation (CD40 ligand plus lymphokines or Stapylococcus aureus protein A) induces no further increase in CstF-64 above that seen for proliferation alone. The rise in CstF-64 is therefore insufficient to induce secretion. After stimulation of a continuously growing B cell line with lymphokines, a switch to Ig secretory mRNA and protein occurs but without a change in the CstF-64 level. Therefore, an increase in CstF-64 levels is not necessary to mediate the differentiation-induced switch to secreted forms of Ig-heavy chain. Because augmentation of CstF-64 levels is neither necessary nor sufficient for Ig secretory mRNA production, we conclude that other lymphokine-induced factors play a role.Treatment of resting cells with agents that caused them to proceed from G 0 to S phase resulted in increased polyadenylation enzyme activity (1-3). An increased rate of polyadenylation of mRNA and accumulation of that newly polyadenylated RNA in the cytoplasm was shown to be a rapid response to entry into S phase, occurring even before large increases in new RNA synthesis (4-6). Many genes have been described whose primary RNA transcripts show a complex pattern of multiple polyadenylation site use in different tissues or growth states (reviewed in ref. 7). The formation of the correct 3Ј-end in some of these RNAs may be regulated as a consequence of changes in polyadenylation factors during proliferation. The recent observation of changes in poly(A) polymerase activity by phosphorylation during the cell cycle (8) reveals one part of this linkage between polyadenylation and cell growth. To determine if the expression of components of the polyadenylation machinery other than poly(A) polymerase may be linked to cell growth, we examined cells that can be induced by the appropriate stimuli to leave G 0 and transition synchronously to S phase, namely serum-starved fibroblasts and resting splenic B cells.Cleavage and polyadenylation of precursor RNAs in vitro requires a number of basal protein factors: cleavage͞ polyadenylation specificity factor (CPSF), CstF, cleavage factors I m and II m and poly(A) polymerase. Synth...