Abstract
Low-density lipoprotein (LDL) receptor-related protein 1B (LRP1B), a member of the LDL receptor family, is frequently inactivated in multiple malignancies including lung cancer. LRP1B is therefore considered as a putative tumor suppressor. Due to its large size (4599 amino acids), until now only minireceptors or receptor fragments have been successfully cloned. To assess the effect of LRP1B on the proliferation of non-small cell lung cancer cells, we constructed and expressed a transfection vector containing the 13.800 bp full-length murine Lrp1b cDNA using a PCR-based cloning strategy. Expression of LRP1B was analyzed by quantitative RT-PCR (qRT-PCR) using primers specific for human LRP1B or mouse Lrp1b. Effective expression of the full length receptor was demonstrated by the appearance of a single 600 kDa band on Western Blots of HEK 293 cells. Overexpression of Lrp1b in non-small cell lung cancer cells with low or absent endogenous LRP1B expression significantly reduced cellular proliferation compared to empty vector-transfected control cells. Conversely, in Calu-1 cells, which express higher endogenous levels of the receptor, siRNA-mediated LRP1B knockdown significantly enhanced cellular proliferation. Taken together, these findings demonstrate that, consistent with the postulated tumor suppressor function, overexpression of full-length Lrp1b leads to impaired cellular proliferation, while LRP1B knockdown has the opposite effect. The recombinant Lrp1b construct represents a valuable tool to unravel the largely unknown physiological role of LRP1B and its potential functions in cancer pathogenesis.
Highlights
LRP1B is a member of the Low-density lipoprotein (LDL) receptor family of lipoprotein receptors, which are involved in different functions in the human body including cholesterol metabolism and atherosclerotic lesion formation
In Calu1 cells, which express higher endogenous levels of the receptor, siRNA-mediated LRP1B knockdown significantly enhanced cellular proliferation. These findings demonstrate that, consistent with the postulated tumor suppressor function, overexpression of full-length Lrp1b leads to impaired cellular proliferation, while LRP1B knockdown has the opposite effect
The construct was introduced by transfection into human cell lines (HEK 293 and non-small cell lung cancer (NSCLC) cell lines)
Summary
LRP1B is a member of the LDL receptor family of lipoprotein receptors, which are involved in different functions in the human body including cholesterol metabolism and atherosclerotic lesion formation. LRP1B was originally discovered during the study of lung cancer cell lines [1]. In nearly 50% of non-small cell lung cancer (NSCLC) cell lines, alterations of the LRP1B gene with homozygous deletions of exons or abnormal transcripts missing portions of the LRP1B sequence were observed. LRP1B was identified as integration site for hepatitis B virus and human papilloma virus presumably with impact on LRP1B expression [16, 17]. Taken together, these observations strongly suggest a role of LRP1B in tumorigenesis and strengthen the original hypothesis of the receptor serving as a tumor suppressor. Expression in smooth muscle cells of the arterial wall has been described [18]
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