Specific recruitment of protein kinase A to the immunoglobulin locus regulates class-switch recombination

Vuong, Bao Q.; Lee, Mieun; Kabir, Shaheen; Irimia, Cristina; Macchiarulo, Stephania; McKnight, G. Stanley; Chaudhuri, Jayanta

doi:10.1038/ni.1708

Cited by 103 publications

(142 citation statements)

References 53 publications

Supporting

Mentioning

134

Contrasting

Order By: Relevance

“…PRKACB and PRKACG obtained from Addgene (Plasmid 20596 and 20597) have been described previously (22). The retroviral vectors pMSCV-IRES-GFP expressing PRKAR2A or R1A were described previously (23).…”

Section: Vectorsmentioning

confidence: 99%

Potassium Channel KCNA1 Modulates Oncogene-Induced Senescence and Transformation

et al. 2013

View full text Add to dashboard Cite

Oncogene-induced senescence (OIS) constitutes a failsafe program that restricts tumor development. However, the mechanisms that link oncogenesis to senescence are not completely understood. We carried out a loss-of-function genetic screen that identified the potassium channel KCNA1 as a determinant of OIS escape that can license tumor growth. Oncogenic stress triggers an increase in KCNA1 expression and its relocation from the cytoplasm to the membrane. Mechanistically, this relocation is due to a loss of protein kinase A (PKA)-induced phosphorylation at residue S446 of KCNA1. Accordingly, sustaining PKA activity or expressing a KCNA1 phosphomimetic mutant maintained KCNA1 in the cytoplasm and caused escape from OIS. KCNA1 relocation to the membrane induced a change in membrane potential that invariably resulted in cellular senescence. Restoring KCNA1 expression in transformation-competent cells triggered variation in membrane potential and blocked RAS-induced transformation, and PKA activation suppressed both effects. Furthermore, KCNA1 expression was reduced in human cancers, and this decrease correlated with an increase in breast cancer aggressiveness. Taken together, our results identify a novel pathway that restricts oncogenesis through a potassium channel-dependent senescence pathway. Cancer Res; 73(16); 5253-65. Ó2013 AACR.

show abstract

Section: Vectorsmentioning

confidence: 99%

Potassium Channel KCNA1 Modulates Oncogene-Induced Senescence and Transformation

et al. 2013

View full text Add to dashboard Cite

show abstract

“…It has been proposed that specific cofactors interact with AID and regulate its activity in a target-specific manner (7). AID has been reported to associate with several proteins including RPA (8,9), protein kinase A (10), RNA polymerase II (Pol II) (11), DNA-PKcs (12), MDM2 (13), Spt6 (14), 14-3-3 (15), Spt5 (16), CTNNBL1 (17), PTBP2 (18), RNA exosome subunits (19), KAP1 (20), and HP1 (20). However, these investigations were mostly focused on CSR, and, thus, the identified factors themselves are not sufficient to explain how AID activity is specifically targeted to the IgV genes during SHM.…”

mentioning

confidence: 99%

Activation-induced cytidine deaminase (AID)-dependent somatic hypermutation requires a splice isoform of the serine/arginine-rich (SR) protein SRSF1

Kanehiro

Todo

Negishi

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Somatic hypermutation (SHM) of Ig variable region (IgV) genes requires both IgV transcription and the enzyme activation-induced cytidine deaminase (AID). Identification of a cofactor responsible for the fact that IgV genes are much more sensitive to AID-induced mutagenesis than other genes is a key question in immunology. Here, we describe an essential role for a splice isoform of the prototypical serine/arginine-rich (SR) protein SRSF1, termed SRSF1-3, in AID-induced SHM in a DT40 chicken B-cell line. Unexpectedly, we found that SHM does not occur in a DT40 line lacking SRSF1-3 (DT40-ASF), although it is readily detectable in parental DT40 cells. Strikingly, overexpression of AID in DT40-ASF cells led to a large increase in nonspecific (off-target) mutations. In contrast, introduction of SRSF1-3, but not SRSF1, into these cells specifically restored SHM without increasing off-target mutations. Furthermore, we found that SRSF1-3 binds preferentially to the IgV gene and inhibits processing of the Ig transcript, providing a mechanism by which SRSF1-3 makes the IgV gene available for AID-dependent SHM. SRSF1 not only acts as an essential splicing factor but also regulates diverse aspects of mRNA metabolism and maintains genome stability. Our findings, thus, define an unexpected and important role for SRSF1, particularly for its splice variant, in enabling AID to function specifically on its natural substrate during SHM.gene conversion | genomic instability S omatic hypermutation (SHM) and class switch recombination (CSR) of Ig genes are key processes needed to generate functional antibodies (Abs) with high affinity in humoral immune responses. Activation-induced cytidine deaminase (AID) is crucial for both SHM and CSR (1, 2), which are initiated by AIDcatalyzed deamination of dC to dU in the variable (V) region and the switch (S) region DNA strands, respectively (3, 4). However, AID has been recently shown to bind numerous chromosomal locations genome-wide and to mutate non-Ig genes, although the mutation frequency of non-Ig genes is much lower than that of the Ig genes (5, 6). It has been proposed that specific cofactors interact with AID and regulate its activity in a target-specific manner (7). AID has been reported to associate with several proteins including RPA (8, 9), protein kinase A (10), RNA polymerase II (Pol II) (11), DNA-PKcs (12), MDM2 (13), Spt6 (14), 14-3-3 (15), Spt5 (16), CTNNBL1 (17), PTBP2 (18), RNA exosome subunits (19), KAP1 (20), and HP1 (20). However, these investigations were mostly focused on CSR, and, thus, the identified factors themselves are not sufficient to explain how AID activity is specifically targeted to the IgV genes during SHM.SHM occurs in IgV region exons starting 100-200 bp downstream of the promoter and extending over 1-2 kb, preferentially targeting WRC/GYW hotspot motifs (3, 4). Importantly, the activity of the SHM machinery strictly depends on transcription of the IgV genes (21, 22). In vitro experiments have shown that replication protein A (RPA) binds to and stabilize...

show abstract

“…AID predominantly localizes in the cytoplasm (9,28) and shuttles between nucleus and cytoplasm. Increasing numbers of proteins have recently been implicated in recruiting AID to the nucleus and to help target it to IgV region and/or S region DNA, including RNA polymerase II (13), RPA (8), and CTNNBL-1 (14) and, most recently, protein kinase A (16). GANP can now be added to this list.…”

Section: Discussionmentioning

confidence: 99%

“…Presumably, AID is recruited to specific Ig regions by a variety of targeting mechanisms that could include cis-acting transcription factors, proteins that associate with AID, and regions of ssDNA formed by transcription bubbles (4,5). There are coimmunoprecipitation (co-IP) data showing interactions between AID and potential recruiter molecules, including RNA polymerase II (13), a spliceosome-associated factor CTNNBL-1 (14), ssDNA-binding protein (RPA) (8,15), and protein kinase A (16). However, it remains unclear how AID is targeted preferentially to actively transcribed Ig genes; nor is it understood how AID is inhibited from deaminating non-Ig transcribed genes (17).…”

mentioning

confidence: 99%

GANP-mediated Recruitment of Activation-induced Cytidine Deaminase to Cell Nuclei and to Immunoglobulin Variable Region DNA

Maeda

Singh

Eda

et al. 2010

Journal of Biological Chemistry

View full text Add to dashboard Cite

AID (activation-induced cytidine deaminase) catalyzes transcription-dependent deamination of C 3 U in immunoglobulin variable (IgV) regions to initiate somatic hypermutation (SHM) in germinal center B-cells. SHM is essential in generating high affinity antibodies. Here we show that when coexpressed with GANP (germinal center-associated nuclear protein) in COS-7 cells, AID is transported from the cytoplasm and concentrated in the nucleus. GANP forms a complex with AID in cotransfected cells in vivo and in vitro. We have isolated AID mutants that bind with reduced affinity to GANP compared with wild type AID. One of these mutants, AID (D143A) binds GANP with a 10-fold lower affinity compared with wild type AID yet retains substantial C-deamination activity in vitro. Mutant AID (D143A) remains localized predominantly in the cytoplasm when coexpressed with GANP. Exogenous expression of GANP in Ramos B-cells promotes binding of AID to IgV DNA and mRNA and increases SHM frequency. These data suggest that GANP may serve as an essential link required to transport AID to B-cell nuclei and to target AID to actively transcribed IgV regions.Affinity maturation of the humoral response proceeds by diversification of Ig genes (1, 2). Diversification requires AID 3 -initiated somatic hypermutation (SHM) within IgV regions and concomitant class switch recombination in Ig switch regions (S regions) (3). AID initiates SHM and class switch recombination by deaminating deoxycytidine residues during transcription of the Ig locus (4, 5). AID deaminates dC 3 dU on single-stranded DNA (ssDNA) and during the transcription of double-stranded DNA (6 -8), presumably on the non-transcribed strand. AID favors deamination in WRC (where W represents A/T, and R is A/G) hot spot motifs (7). The observation of strongly enhanced C 3 T mutations in variable regions and S regions, which conform to the mutational signature of AID (7), offers convincing evidence that AID is acting directly on the transcribed DNA. AID is located predominantly in the cytoplasm of activated B-cells (9) and has a nuclear localization signal motif in its N-terminal region and a nuclear export signal at its C terminus (10 -12). Deletion of the nuclear export signal results in nuclear accumulation (11, 12) but does not result in an increase in mutations in S regions (11). There must be a means to regulate the import of AID into the nucleus and then to direct its access to transcribed Ig loci. Presumably, AID is recruited to specific Ig regions by a variety of targeting mechanisms that could include cis-acting transcription factors, proteins that associate with AID, and regions of ssDNA formed by transcription bubbles (4, 5). There are coimmunoprecipitation (co-IP) data showing interactions between AID and potential recruiter molecules, including RNA polymerase II (13), a spliceosome-associated factor CTNNBL-1 (14), ssDNA-binding protein (RPA) (8, 15), and protein kinase A (16). However, it remains unclear how AID is targeted preferentially to actively transcribed Ig genes; no...

show abstract

Specific recruitment of protein kinase A to the immunoglobulin locus regulates class-switch recombination

Cited by 103 publications

References 53 publications

Potassium Channel KCNA1 Modulates Oncogene-Induced Senescence and Transformation

Potassium Channel KCNA1 Modulates Oncogene-Induced Senescence and Transformation

Activation-induced cytidine deaminase (AID)-dependent somatic hypermutation requires a splice isoform of the serine/arginine-rich (SR) protein SRSF1

GANP-mediated Recruitment of Activation-induced Cytidine Deaminase to Cell Nuclei and to Immunoglobulin Variable Region DNA

Contact Info

Product

Resources

About