Burkitt lymphoma (BL) is a highly malignant non-Hodgkin's lymphoma that is closely
related to the abnormal expression of genes. Familial acute myelogenous leukemia
related factor (FAMLF; GenBank accession No. EF413001.1) is a novel
gene that was cloned by our research group, and miR-181b is located in the intron of
the FAMLF gene. To verify the role of miR-181b and
FAMLF in BL, RNAhybrid software was used to predict target site
of miR-181b on FAMLF and real-time quantitative PCR (RQ-PCR) was
used to detect expression of miR-181b and FAMLF in BL patients, Raji
cells and unaffected individuals. miR-181b was then transfected into Raji and CA46
cell lines and FAMLF expression was examined by RQ-PCR and western
blotting. Further, Raji cells viability and proliferation were detected by MTT and
clone formation, and Raji cell cycle and apoptosis were detected by flow cytometry.
The results showed that miR-181b can bind to bases 21–42 of the
FAMLF 5′ untranslated region (UTR), FAMLF was
highly expressed and miR-181b was lowly expressed in BL patients compared with
unaffected individuals. FAMLF expression was significantly and
inversely correlated to miR-181b expression, and miR-181b negatively regulated
FAMLF at posttranscriptional and translational levels. A
dual-luciferase reporter gene assay identified that the 5′ UTR of
FAMLF mRNA contained putative binding sites for miR-181b.
Down-regulation of FAMLF by miR-181b arrested cell cycle, inhibited
cell viability and proliferation in a BL cell line model. Our findings explain a new
mechanism of BL pathogenesis and may also have implications in the therapy of
FAMLF-overexpressing BL.
The familial acute myeloid leukemia related factor gene (FAMLF) was
previously identified from a familial AML subtractive cDNA library and shown to
undergo alternative splicing. This study used real-time quantitative PCR to
investigate the expression of the FAMLF alternative-splicing
transcript consensus sequence (FAMLF-CS) in peripheral blood
mononuclear cells (PBMCs) from 119 patients with de novo acute
leukemia (AL) and 104 healthy controls, as well as in CD34+cells from 12
AL patients and 10 healthy donors. A 429-bp fragment from a novel splicing variant of
FAMLF was obtained, and a 363-bp consensus sequence was targeted
to quantify total FAMLF expression. Kruskal-Wallis, Nemenyi,
Spearman's correlation, and Mann-Whitney U-tests were used to analyze the data.
FAMLF-CS expression in PBMCs from AL patients and
CD34+ cells from AL patients and controls was significantly higher than
in control PBMCs (P<0.0001). Moreover,FAMLF-CS expression in
PBMCs from the AML group was positively correlated with red blood cell count
(rs
=0.317, P=0.006), hemoglobin levels (rs
=0.210, P=0.049), and percentage of peripheral blood blasts
(rs
=0.256, P=0.027), but inversely correlated with hemoglobin levels in
the control group (rs
=–0.391, P<0.0001). AML patients with high CD34+
expression showed significantly higherFAMLF-CS expression than those
with low CD34+ expression (P=0.041). Our results showed
thatFAMLF is highly expressed in both normal and malignant
immature hematopoietic cells, but that expression is lower in normal mature
PBMCs.
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