American Society of Hematology

Figure 1.

$Figure 1. Effect of BCR/ABL on HNRPK mRNA and protein expression. (Ai) Northern blot shows expression of Hnrpk mRNA in parental, untreated, and imatinib-treated BCR/ABL-expressing myeloid progenitor 32Dcl3 cells. (Aii) RT-PCR on nuclear RNA shows levels of Hnrpk pre-mRNA in parental and BCR/ABL-expressing myeloid progenitor 32Dcl3 cells (+RT). The amplified PCR product is comprehensive of part of exon 2, intron 2, and part of exon 3 of the mouse Hnrpk gene.-RT indicates that PCR was performed on non-reverse-transcribed nuclear RNA. GRB2 mRNA levels were measured as control. (Aiii) Effect of actinomycin D on Hnrpk mRNA levels in parental and BCR/ABL-expressing 32Dcl3 cells. Cells were incubated for the indicated time with actinomycin D used at a concentration (5 μg/mL) that reportedly inhibits RNA polII-dependent transcription. (Bi) Western blots show HNRPK protein levels in parental 32Dcl3 cells (lane 1) and in untreated (lane 2) and imatinib-treated (lane 3) 32D-BCR/ABL cells, in BCR/ABL-induced (doxycycline 1 mg/mL, 24 hours) TonB210.1 lymphoid progenitors (lanes 4-5), and in untreated and imatinib-treated Ph1 K562 cells (lanes 6-7). (C) HNRPK protein stability was determined by anti-HNRPK Western blots in untreated and imatinib-treated 32D-BCR/ABL cells exposed to the protein synthesis inhibitor cycloheximide (CHX). GRB2 levels were monitored as control for equal loading. (Di) Plot shows HNRPK protein levels expressed as log of arbitrary densitometric units after normalization with GRB2 levels in mononuclear cells from CML patients in the chronic (CML-CP) and blast crisis (CML-BC) phases. (P = .001; Wicoxon rank sum test). (Dii) Graphs shows GRB2-normalized HNRPK protein levels expressed as densitometric units in the CD34+ fraction of bone marrow from 2 healthy donors (NBM), and 2 CML-CP, 1 CML-AP, and 2 CML-BC patients. The Western blot in the inset shows total tyrosine phosphorylation, and the arrow indicates levels of active BCR/ABL. (Diii) HNRPK protein expression in untreated and imatinib-treated CD34+ CML-BC patient marrow cells. GRB2 was used as control for equal loading.$

Effect of BCR/ABL on HNRPK mRNA and protein expression. (Ai) Northern blot shows expression of Hnrpk mRNA in parental, untreated, and imatinib-treated BCR/ABL-expressing myeloid progenitor 32Dcl3 cells. (Aii) RT-PCR on nuclear RNA shows levels of Hnrpk pre-mRNA in parental and BCR/ABL-expressing myeloid progenitor 32Dcl3 cells (+RT). The amplified PCR product is comprehensive of part of exon 2, intron 2, and part of exon 3 of the mouse Hnrpk gene.-RT indicates that PCR was performed on non-reverse-transcribed nuclear RNA. GRB2 mRNA levels were measured as control. (Aiii) Effect of actinomycin D on Hnrpk mRNA levels in parental and BCR/ABL-expressing 32Dcl3 cells. Cells were incubated for the indicated time with actinomycin D used at a concentration (5 μg/mL) that reportedly inhibits RNA polII-dependent transcription. (Bi) Western blots show HNRPK protein levels in parental 32Dcl3 cells (lane 1) and in untreated (lane 2) and imatinib-treated (lane 3) 32D-BCR/ABL cells, in BCR/ABL-induced (doxycycline 1 mg/mL, 24 hours) TonB210.1 lymphoid progenitors (lanes 4-5), and in untreated and imatinib-treated Ph1 K562 cells (lanes 6-7). (C) HNRPK protein stability was determined by anti-HNRPK Western blots in untreated and imatinib-treated 32D-BCR/ABL cells exposed to the protein synthesis inhibitor cycloheximide (CHX). GRB2 levels were monitored as control for equal loading. (Di) Plot shows HNRPK protein levels expressed as log of arbitrary densitometric units after normalization with GRB2 levels in mononuclear cells from CML patients in the chronic (CML-CP) and blast crisis (CML-BC) phases. (P = .001; Wicoxon rank sum test). (Dii) Graphs shows GRB2-normalized HNRPK protein levels expressed as densitometric units in the CD34⁺ fraction of bone marrow from 2 healthy donors (NBM), and 2 CML-CP, 1 CML-AP, and 2 CML-BC patients. The Western blot in the inset shows total tyrosine phosphorylation, and the arrow indicates levels of active BCR/ABL. (Diii) HNRPK protein expression in untreated and imatinib-treated CD34⁺ CML-BC patient marrow cells. GRB2 was used as control for equal loading.

This Feature Is Available To Subscribers Only