1: Mol Cell Biol. 2005 Jul;25(14):6021-30. 

Sites that direct nuclear compartmentalization are near the 5' end of the mouse
immunoglobulin heavy-chain locus.

Yang Q, Riblet R, Schildkraut CL.

Department of Cell Biology (CH 416), Albert Einstein College of Medicine, 1300
Morris Park Ave., Bronx, New York 10461, USA.

VDJ rearrangement in the mouse immunoglobulin heavy chain (Igh) locus involves a
combination of events, including a large change in its nuclear
compartmentalization. Prior to rearrangement, Igh moves from its default
peripheral location near the nuclear envelope to an interior compartment, and
after rearrangement it returns to the periphery. To identify any sites in Igh
responsible for its association with the periphery, we systematically analyzed
the nuclear positions of the Igh locus in mouse non-B- and B-cell lines and,
importantly, in primary splenic lipopolysaccharide-stimulated B cells and
plasmablasts. We found that a broad approximately 1-Mb region in the 5' half of
the variable-gene region heavy-chain (Vh) locus regularly colocalizes with the
nuclear lamina. The 3' half of the Vh gene region is less frequently colocalized
with the periphery, while sequences flanking the Vh gene region are infrequently
so. Importantly, in plasmacytomas, VDJ rearrangements that delete most of the Vh
locus, including part of the 5' half of the Vh gene region, result in loss of
peripheral compartmentalization, while deletion of only the proximal half of the
Vh gene region does not. In addition, when Igh-Myc translocations move the Vh
genes to a new chromosome, the distal Vh gene region is still associated with
the nuclear periphery. Thus, the Igh region that interacts with the nuclear
periphery is localized but is likely comprised of multiple sites that are
distributed over approximately 1 Mb in the 5' half of the Vh gene region. This
5' Vh gene region that produces peripheral compartmentalization is the same
region that is distinguished by requirements for interleukin-7, Pax5, and Ezh2
for rearrangement of the Vh genes.

PMID: 15988016 [PubMed - indexed for MEDLINE]


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2: Leukemia. 2004 Dec;18(12):2026-31. 

Rapid amplification of immunoglobulin heavy chain switch (IGHS) translocation
breakpoints using long-distance inverse PCR.

Sonoki T, Willis TG, Oscier DG, Karran EL, Siebert R, Dyer MJ.

Academic Haematology and Cytogenetics, Institute of Cancer Research, UK.

Molecular cloning of immunoglobulin heavy chain (IGH) translocation breakpoints
identifies genes of biological importance in the development of normal and
malignant B cells. Long-distance inverse PCR (LDI-PCR) was first applied to
amplification of IGH gene translocations targeted to the joining (IGHJ) regions.
We report here successful amplification of the breakpoint of IGH translocations
targeted to switch (IGHS) regions by LDI-PCR. To detect IGHS translocations,
Southern blot assays using 5' and 3' switch probes were performed. Illegitimate
Smu rearrangements were amplified from the 5' end (5'Smu LDI-PCR) from the
alternative derivative chromosome, and those of Sgamma or Salpha were amplified
from the 3' end (3'Sgamma or 3'alpha LDI-PCR) from the derivative chromosome 14.
Using a combination of these methods, we have succeeded in amplifying IGHS
translocation breakpoints involving FGFR3/MMSET on 4p16, BCL6 on 3q27, MYC on
8q24, IRTA1 on 1q21 and PAX5 on 9p13 as well as BCL11A on 2p13 and CCND3 on
6p21. The combination of LDI-PCR for IGHJ and IGHS allows rapid molecular
cloning of almost all IGH gene translocation breakpoints.

PMID: 15496980 [PubMed - indexed for MEDLINE]


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3: J Exp Med. 2004 Jun 21;199(12):1689-700. 

Early B cell factor promotes B lymphopoiesis with reduced interleukin 7
responsiveness in the absence of E2A.

Seet CS, Brumbaugh RL, Kee BL.

Department of Pathology, University of Chicago, 5841 S. Maryland Avenue, MC
1089, Chicago, IL 60637, USA.

The basic helix-loop-helix transcription factors encoded by the E2A gene
function at the apex of a transcriptional hierarchy involving E2A, early B cell
factor (EBF), and Pax5, which is essential for B lymphopoiesis. In committed B
lineage progenitors, E2A proteins have also been shown to regulate many
lineage-associated genes. Herein, we demonstrate that the block in B
lymphopoiesis imposed by the absence of E2A can be overcome by expression of
EBF, but not Pax5, indicating that EBF is the essential target of E2A required
for development of B lineage progenitors. Our data demonstrate that EBF, in
synergy with low levels of alternative E2A-related proteins (E proteins), is
sufficient to promote expression of most B lineage genes. Remarkably, however,
we find that E2A proteins are required for interleukin 7-dependent proliferation
due, in part, to a role for E2A in optimal expression of N-myc. Therefore, high
levels of E protein activity are essential for the activation of EBF and N-myc,
whereas lower levels of E protein activity, in synergy with other B lineage
transcription factors, are sufficient for expression of most B lineage genes.

PMID: 15210745 [PubMed - indexed for MEDLINE]


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4: Leuk Lymphoma. 2003;44 Suppl 3:S21-6. 

Pathobiology of primary mediastinal B-cell lymphoma.

Pileri SA, Zinzani PL, Gaidano G, Falini B, Gaulard P, Zucca E, Sabattini E,
Ascani S, Rossi M, Cavalli F; International Extranodal Lymphoma Study Group.

Institute of Haematology Seragnoli, Bologna University, Policinico S. Orsola,
via Massarenti 9, 40138, Bologna, Italy. pileri@med.unibo.it

Controversy still exists over the response to therapy and prognosis of patients
with primary mediastinal B-cell lymphoma (PMBL). Recent data from the
International Extranodal Lymphoma Study Group (IELSG) suggest that a MACOP-B
(methotrexate, adriamycin, cyclophosphamide, vincristine, prednisone, bleomycin)
chemotherapy regimen followed by radiotherapy may be a better induction strategy
than other previously used treatments. Although the pathobiology of PMBL has
been widely studied, its precise histology, phenotype, and molecular
characteristics are still not clear. To date, phenotypic analysis has revealed
the following phenotype: positivity for CD45 and CD20, but negativity for CD3,
CD10, CD21, Class I/II major histocompatibility antigens, and a variety of other
immunohistochemical markers. CD79a is generally detected, despite an absence of
surface immunoglobulins (Igs). CD30 staining is observed in most cases, but is
weaker and less homogeneous than in classic Hodgkin's lymphoma or anaplastic
large cell lymphoma. BCL-2 protein is usually expressed but there are few data
describing the expression of MUM1/IRF4, PAX5/BSAP, BCL-6, or the B-cell
transcription factors BOB.1, Oct-2, and PU.1. Cytogenetic studies reveal gains
in segments of chromosome 9p, including amplification of the REL proto-oncogene
and the tyrosine kinase gene JAK2. Other molecular findings include: C-myc
mutations or rearrangements, p53 mutations, IgV(H), gene mutations, and bcl-2
and mal over-expression. bcl-6 mutations and bcl-2 gene rearrangements are
generally absent, suggesting that PMBL is of pre-germinal center (GC) origin.
However, two recent reports show isotype-switched Ig genes with a high frequency
of somatic hypermutations as well as variants in the 5' noncoding region of the
bcl-6 gene. The IELSG collected 137 PMBL cases for extensive pathologic review.
Histologically, the lymphomatous growth was predominantly diffuse with sclerosis
that induced compartmentalized cell aggregation. It consisted of large cells
with varying degrees of nuclear polymorphism and clear to basophilic cytoplasm.
Molecular analysis was performed on 40 cases and showed novel findings. More
than half of the cases displayed bcl-6 gene mutations, which usually occurred
together with functioning somatic IgV(H) gene mutations, and BCL-6 and/or
MUM1/IRF4 expression. The present study supports the concept that PBML is
derived from activated GC or post-germinal center cells. However, it differs
from other aggressive B-cell lymphomas in that it shows defective Ig production
despite the expression of Oct-2, BOB.1, and PU.1 transcription factors, and a
lack of IgV(H) gene crippling mutations.

Publication Types:
    Review
    Review, Tutorial

PMID: 15202521 [PubMed - indexed for MEDLINE]


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5: Blood. 2004 Mar 1;103(5):1869-75. Epub 2003 Oct 30. 

Primary diffuse large B-cell lymphomas of the central nervous system are
targeted by aberrant somatic hypermutation.

Montesinos-Rongen M, Van Roost D, Schaller C, Wiestler OD, Deckert M.

Department of Neuropathology, University of Cologne, Koln, Germany.

We have addressed whether aberrant ongoing hypermutation can be detected in the
proto-oncogenes PIM1, c-MYC, RhoH/TTF, PAX5, and the tumor-suppressor gene CD95
in primary central nervous system lymphomas (PCNSLs) derived from
immunocompetent HIV-negative patients. Nine of 10 PCNSLs analyzed harbored
somatic mutations in the PIM1, c-MYC, RhoH/TTF, and PAX5 genes, but not in the
CD95 gene, with 8 tumors carrying alterations in at least 2 of these genes.
Furthermore, ongoing aberrant mutation was evidenced in a subset of PCNSLs (2 of
3). Although most of the mutations corresponded to base pair substitutions,
deletions were also present. The mean mutation frequency was approximately
60-fold lower for these genes compared with the values obtained for
immunoglobulin genes in PCNSL. They were increased 2- to 5-fold compared with
extracerebral diffuse large B-cell lymphoma (DLBCL). In summary, our data
demonstrate aberrant somatic hypermutations at high frequency in the PIM1, PAX5,
RhoH/TTF, and c-MYC genes in most PCNSLs. These findings may indicate a
pathogenic role for aberrant somatic hypermutation in PCNSL development. In
contrast, although mutations were detected in exon 9 of the CD95 gene, the lack
of mutations in the 5' region provides no evidence for the CD95 gene as a target
for aberrant somatic mutation.

PMID: 14592832 [PubMed - indexed for MEDLINE]


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6: Blood. 2003 Mar 1;101(5):1950-5. Epub 2002 Oct 24. 

Oscillation between B-lymphoid and myeloid lineages in Myc-induced hematopoietic
tumors following spontaneous silencing/reactivation of the EBF/Pax5 pathway.

Yu D, Allman D, Goldschmidt MH, Atchison ML, Monroe JG, Thomas-Tikhonenko A.

Departments of Pathobiology, Pathology and Laboratory Medicine, and Animal
Biology, University of Pennsylvania, Philadelphia, PA 19104-6051, USA.

B lymphomagenesis is an uncontrolled expansion of immature precursors that fail
to complete their differentiation program. This failure could be at least partly
explained by inappropriate expression of several oncogenic transcription
factors, such as Pax5 and Myc. Both Pax5 and c-Myc are implicated in the
pathogenesis of non-Hodgkin lymphomas. To address their role in lymphomagenesis,
we analyzed B-cell lymphomas derived from p53-null bone marrow progenitors
infected in vivo by a Myc-encoding retrovirus. All Myc-induced lymphomas
invariably maintained expression of Pax5, which is thought to be incompatible
with terminal differentiation. However, upon culturing in vitro, several cell
lines spontaneously down-regulated Pax5 and its target genes CD19, N-Myc, and
MB1. Unexpectedly, other B-cell markers (eg, CD45R) were also down-regulated,
and markers of myeloid lineage (CD11b and F4/80 antigen) were acquired instead.
Moreover, cells assumed the morphology reminiscent of myeloid cells. A pool of
F4/80-positive cells as well as several single-cell clones were obtained and
reinjected into syngeneic mice. Remarkably, pooled cells rapidly re-expressed
Pax5 and formed tumors of relatively mature lymphoid phenotype, with surface
immunoglobulins being abundantly expressed. Approximately half of tumorigenic
single-cell clones also abandoned myeloid differentiation and gave rise to B
lymphomas. However, when secondary lymphoma cells were returned to in vitro
conditions, they once again switched to myeloid differentiation. This process
could be curbed via enforced expression of retrovirally encoded Pax5. Our data
demonstrate that some Myc target cells are bipotent B-lymphoid/myeloid
progenitors with the astonishing capacity to undergo successive rounds of
lineage switching.

PMID: 12406913 [PubMed - indexed for MEDLINE]


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7: Nature. 2001 Jul 19;412(6844):341-6. 

Hypermutation of multiple proto-oncogenes in B-cell diffuse large-cell
lymphomas.

Pasqualucci L, Neumeister P, Goossens T, Nanjangud G, Chaganti RS, Kuppers R,
Dalla-Favera R.

Institute for Cancer Genetics and the Department of Pathology, Columbia
University, New York, New York 10032, USA.

Genomic instability promotes tumorigenesis and can occur through various
mechanisms, including defective segregation of chromosomes or inactivation of
DNA mismatch repair. Although B-cell lymphomas are associated with chromosomal
translocations that deregulate oncogene expression, a mechanism for genome-wide
instability during lymphomagenesis has not been described. During B-cell
development, the immunoglobulin variable (V) region genes are subject to somatic
hypermutation in germinal-centre B cells. Here we report that an aberrant
hypermutation activity targets multiple loci, including the proto-oncogenes
PIM1, MYC, RhoH/TTF (ARHH) and PAX5, in more than 50% of diffuse large-cell
lymphomas (DLCLs), which are tumours derived from germinal centres. Mutations
are distributed in the 5' untranslated or coding sequences, are independent of
chromosomal translocations, and share features typical of V-region-associated
somatic hypermutation. In contrast to mutations in V regions, however, these
mutations are not detectable in normal germinal-centre B cells or in other
germinal-centre-derived lymphomas, suggesting a DLCL-associated malfunction of
somatic hypermutation. Intriguingly, the four hypermutable genes are susceptible
to chromosomal translocations in the same region, consistent with a role for
hypermutation in generating translocations by DNA double-strand breaks. By
mutating multiple genes, and possibly by favouring chromosomal translocations,
aberrant hypermutation may represent the major contributor to lymphomagenesis.

PMID: 11460166 [PubMed - indexed for MEDLINE]


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8: Int Rev Immunol. 2001 Feb;20(1):65-82. 

Pax5 determines the identity of B cells from the beginning to the end of
B-lymphopoiesis.

Nutt SL, Eberhard D, Horcher M, Rolink AG, Busslinger M.

Research Institute of Molecular Pathology, Dr. Bohr-Gasse 7, A-1030 Vienna,
Austria.

Despite being one of the most intensively studied cell types, the molecular
basis of B cell specification is largely unknown. The Pax5 gene encoding the
transcription factor BSAP is required for progression of B-lymphopoiesis beyond
the pro-B cell stage. Pax5-deficient pro-B cells are, however, not yet committed
to the B-lymphoid lineage, but instead have a broad lymphomyeloid developmental
potential. Pax5 appears to mediate B-lineage commitment by repressing the
transcription of non-B-lymphoid genes and by simultaneously activating the
expression of B-lineage-specific genes. Pax5 thus functions both as a
transcriptional repressor and activator, depending on its interactions with
corepressors of the Groucho protein family or with positive regulators such as
the TATA-binding protein. Once committed to the B-lineage, B cells require Pax5
function to maintain their B-lymphoid identity throughout B cell development.

Publication Types:
    Review
    Review, Tutorial

PMID: 11342298 [PubMed - indexed for MEDLINE]


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9: Clin Cancer Res. 1995 Feb;1(2):207-14. 

PAX5 expression correlates with increasing malignancy in human astrocytomas.

Stuart ET, Kioussi C, Aguzzi A, Gruss P.

Department of Molecular Cell Biology, Max-Planck Institute for Biophysical
Chemistry, Am Fassberg, 37077 Gottingen, Germany.

Rearrangements concerning chromosome 9p are a late event in the progression of
human astrocytic tumors to their most malignant form. The expression of PAX5,
which maps to chromosome 9p13, was studied in primary human brain tumors of
astrocytic origin. Whereas murine Pax5 is not expressed in the forebrain at any
stage, PAX5 expression was increased in a range of astrocytomas (WHO grades
II-IV) which originated in the forebrain. Expression of PAX5 was limited to
those cells which also expressed the oncogenes myc, fos, or jun singularly or in
combination. The epidermal growth factor receptor was highly expressed in
glioblastoma multiform tumors in areas which were also highly PAX5 positive. We
conclude that the missappropriate expression of PAX5 may aid in promoting the
progression of astrocytomal malignancy.

PMID: 9815975 [PubMed - indexed for MEDLINE]


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