Physical mapping, amplification, and overexpression of the mouse mdr gene family in multidrug-resistant cells.

Abstract

The mouse mdr gene family consists of three distinct genes (mdrl, mdr2, and mdr3), for which we have isolated full-length cDNA clones. cDNA subfragments corresponding to discrete regions showing little sequence conservation among the three mdr genes were used as gene-specific DNA probes in hybridization experiments. Long-range mapping by pulse-field gel electrophoresis indicated that the three mdr genes are closely linked on a genomic DNA segment of approximately 625 kilobases. The gene order and direction of transcription of the three genes were determined and indicate the arrangement (5') mdr3 (3')-(5') mdrl (3')-(3') mdr2 (5'). Southern blotting analyses of genomic DNA from a panel of independently derived multidrug-resistant cell lines identified mdr gene amplification in 10 of 12 cell lines studied. In individual cell lines showing gene amplification, the copy number of each of the three mdr genes was identical, suggesting that the three mdr genes became amplified as part of a single amplicon in these cells. Although increased expression of all three mdr genes was detected in 2 of 12 cell lines tested, multidrug resistance was associated in 10 of 12 lines with the independent overexpression of either mdrl (7 of 12) or mdr3 (3 of 12) but not mdr2. mdrl overexpression was consistently associated with gene amplification, while increased mdr3 expression was detected in certain cell lines that did not show gene amplification. Increased levels of mdrl mRNA were linked to the overexpression of a P glycoprotein of apparent molecular weight 180,000 to 200,000, whereas increased mdr3 expression resulted in increased expression of a P glycoprotein of molecular weight 160,000 to 180,000. Our results suggest that at least two members of the mouse mdr gene family, mdrl and mdr3, can independently confer multidrug resistance in the cell lines examined.In cultured cells, multidrug resistance is associated with the overexpression of a heterogeneous group of antigenically related membrane phosphoglycoproteins, termed P glycoproteins (12). P glycoproteins are encoded by a small family of closely related genes given the appellation pgp or mdr which often become amplified in highly drug-resistant cell lines (reviewed in references 8, 12, and 15). The mdrlpgp gene family is composed of three members in rodents (hamster and mouse) and two members in humans (24 suggest that P glycoprotein functions in drug-resistant cells as an ATP-dependent drug efflux pump.Sequence analysis of full-length cDNA clones encoding mouse mdrl, mdr2, and mdr3 indicates that the three encoded proteins are highly homologous (overall amino acid homology of 75 to 85%) and share the same predicted structural domains (10a, 19, 20

Genomic Sequence Analysis of the Mouse Naip Gene Array

“…Examples of closely linked gene families whose members have divergences in function are seen in the cases of the color-vision genes and the betaglobins (Nathans et al 1986;Yokoyama et al 1993;Fritsch et al 1980;Hardies et al 1984). Similarly, there are examples of the occurrence of closely linked gene copies that are redundant in function, such as is seen in the observed amplification of ribosomal RNA genes in various organisms and in the cellular aquisition of resistance to chemotherapeutic agents (Nath and Bollon 1977;Raymond et al 1990). …”

Calorie Restriction Increases P-Glycoprotein and Decreases Intestinal Absorption of Digoxin in Mice

“…P-gp transports a wide range of structurally different therapeutics including colchicine, tacrolimus, quinidine, chemotherapeutic drugs (such as etoposide, doxorubicin, and vinblastine), cardiac glycosides (such as digoxin), glucocorticoids (such as dexamethasone), and human immunodeficiency virus type 1 antiretroviral therapy drugs (Aller et al, 2009). Overexpression of Abcb1a and -1b can confer drug resistance by increasing efflux of these drugs from cells (Dhir et al, 1990;Raymond et al, 1990) and is a well-established cause of resistance to cancer chemotherapeutic drugs. Abcb1a is mainly expressed in the gastrointestinal tract, testis, and capillaries within the brain (Cui et al, 2009).…”

Defect of multidrug-resistance 3 gene expression in a subtype of progressive familial intrahepatic cholestasis

“…[27][28][29][30] However, no associated genetic defect has been identified. It has The mouse mdr2 gene (standard gene symbol Pgy-been suggested that one subtype of PFIC may be attrib-3) and its human homolog MDR3 (also called MDR2, utable to a defect in primary bile acid secretion, 29 and it standard gene symbol PGY3) are members of the multi-has been established that children affected by another drug gene family [1][2][3][4][5][6][7] that do not confer multidrug resis-subtype have a defect in primary bile acid synthesis. 30 tance [8][9][10][11] but encode P-glycoproteins, which act as phos-In both of these subtypes, cholestasis is associated with phatidylcholine flippases.…”