Identification of different Ikaros cDNA transcripts in Philadelphia-positive adult acute lymphoblastic leukemia by a high-throughput capillary electrophoresis sizing method.
BACKGROUND
Ikaros is the prototypic member of a Kruppel-like zinc finger transcription factor subfamily that is required for normal hematopoietic cell differentiation and proliferation, particularly in the lymphoid lineages. Alternative splicing can generate multiple Ikaros isoforms that lack different numbers of exons and have different functions. Shorter isoforms, which lack the amino-terminal domain that mediates sequence-specific DNA binding, exert a dominant negative effect and inhibit the ability of longer heterodimer partners to bind DNA.


METHODS
In this study, we developed a high-throughput capillary electrophoresis sizing method to detect and quantify different Ikaros cDNA transcripts.


RESULTS
We demonstrated that Philadelphia chromosome-positive acute lymphoblastic leukemia cells expressed high levels of the non-DNA-binding isoform Ik6 that was generated following IKZF1 genomic deletions (19/46 patients, 41%). Furthermore, a recurring 60 bp insertion immediately upstream of exon 5, at the exon 3/exon 5 junction, was frequently detected in the Ik2 and Ik4 isoforms. This insertion occurred either alone or together with an in-frame ten amino acid deletion that was due to a 30 bp loss at the end of exon 7. Both the alterations are due to the selection of alternative cryptic splice sites and have been suggested to cause impaired DNA-binding activity. Non-DNA-binding isoforms were localized in the cytoplasm whereas the DNA-binding isoforms were localized in the nucleus.


CONCLUSIONS
Our findings demonstrate that both aberrant splicing and genomic deletion leading to different non-DNA-binding Ikaros cDNA transcripts are common features of Philadelphia chromosome-positive acute lymphoblastic leukemia.
Iacobucci
Ilaria
I
Department of Hematology/Oncology L. and A. Seràgnoli S.Orsola Malpighi Hospital, University of Bologna, Bologna, Italy.
Lonetti
Annalisa
A
Cilloni
Daniela
D
Messa
Francesca
F
Ferrari
Anna
A
Zuntini
Roberta
R
Ferrari
Simona
S
Ottaviani
Emanuela
E
Arruga
Francesca
F
Paolini
Stefania
S
Papayannidis
Cristina
C
Piccaluga
Pier Paolo
PP
Soverini
Simona
S
Saglio
Giuseppe
G
Pane
Fabrizio
F
Baruzzi
Anna
A
Vignetti
Marco
M
Berton
Giorgio
G
Vitale
Antonella
A
Chiaretti
Sabina
S
Müschen
Markus
M
Foà
Robin
R
Baccarani
Michele
M
Martinelli
Giovanni
G
D000293:Adolescent; D000328:Adult; D000368:Aged; D018076:DNA, Complementary / Q000032:analysis; D019075:Electrophoresis, Capillary / Q000379:methods*; D006801:Humans; D051740:Ikaros Transcription Factor / Q000032:analysis / Q000235:genetics*; D008875:Middle Aged; D054198:Precursor Cell Lymphoblastic Leukemia-Lymphoma / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis; D055815:Young Adult
D018076:DNA, Complementary; D012333:RNA, Messenger; D051740:Ikaros Transcription Factor