Decreased E12 and/or E47 Transcription Factor Activity in the Bone Marrow As Well As in the Spleen of Aged Mice

Abstract

The E2A-encoded transcription factors E12 and E47 are key regulators of B cell functions. They bind to the E-box site, found in regulatory regions of B cell-specific genes; promote cell survival of early pre-B cells; help to initiate Ig rearrangements; and are also involved in class switch in mature B cells in the periphery. We have investigated the expression and function of E47 and E12 in IL-7-expanded pro-B/pre-B cell precursors and in unstimulated or LPS-activated splenic B cells from young and old BALB/c mice. Results show that B cell precursors from the bone marrow of old mice exhibit a reduced expression of E2A proteins and a reduced ability to bind DNA, as compared with young mice. In the spleen, E2A protein expression and DNA binding are present in unstimulated B cells from young mice and, to a significantly lesser extent, from old mice. These are both strongly induced by activation in splenic B cells from young mice but only moderately induced in old mice, indicating that aging affects the expression and activity of E2A-encoded genes and also that DNA binding correlates with the amount of protein expression. The levels of E2A DNA binding in the spleen correlate with those in the bone marrow for individual mice. In splenic mature B cells, only E47/E47 complexes bind DNA; whereas in bone marrow B cell precursors, E47/E12 complexes participate in DNA binding. Only nuclear extracts of splenic mature B cells, but both nuclear and cytoplasmic extracts of bone marrow B cell precursors, exhibit DNA binding.

Accelerated Notch-Dependent Degradation of E47 Proteins in Aged B Cell Precursors Is Associated with Increased ERK MAPK Activation

“…In aged mice, as we have shown herein and elsewhere (3,9,11,12), pro-B/early pre-B cells have reduced overall levels of E47 protein. Although our previous studies have relied on Western blotting of lysates from pooled B cell precursors, the current study using fluorescence flow cytometry provides for determination of E47 protein expression by individual pro-B/early pre-B cells in aged mice.…”

“…Previously, we demonstrated that E2A-encoded proteins were reduced in B cell precursors derived from aged BALB/c bone marrow upon culture with recombinant mouse IL-7 (9,11,12). This reflected increased protein turnover rather than alterations in expression of E2A-encoded mRNA (12).…”

“…Both transcription of the RAG enzymes and the surrogate L chain proteins in B cell precursors are, in part, mediated by the E2A-encoded transcription factors E47 and E12 (10). Previously, we have shown that E2A-encoded protein levels are reduced in B cell precursors from aged mice after their expansion with IL-7 in vitro (9,11,12). This was associated with reduction in the expression of 5 protein and reflected in decreased 5 and VpreB mRNAs (9).…”

Aging murine B cells have decreased class switch induced by anti-CD40 or BAFF

“…BAFF-R forward, CCC-AGA-CTC-GGA-ACT-GTC-CC; BAFF-R reverse, GTA-GAG-ATC-CCT-GGG-TTC-CAG; TACI forward, CAG-GTA-CTA-CGA-CCA-TCT-CCT-G; TACI reverse, TCG-TCG-CAA-GCC-TCT-GTC-AC; BCMA forward, CCC-ACA-GAG-AAT-CTG-CTG-TTC-TTC; BCMA reverse, AAA-GAT-CCT-GTC-GTC-ACC-AGC-CC. E2A primers amplify both E12 and E47, but E47 is the major E2A-encoded mRNA and protein in splenic B cells (Frasca et al, 2003). Subsequent experiments using E47 primers show similar results.…”

“…We have shown (Frasca et al, 2004a) that splenic B cells from old mice have reduced levels of E47, and subsequently AID, leading to poor CSR and production of secondary isotypes in senescence. We have previously shown (Frasca et al, 2003) that in splenic mature B cells, only E47/E47 homodimeric complexes bind DNA, whereas in bone marrow B cell precursors, E47/E12 complexes participate in DNA binding. E12 is present at very low levels in activated splenic B cells from young mice and is almost undetectable in old B cells.…”