Analysis of mutant DNA polymerase gamma in patients with mitochondrial DNA depletion.
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Analysis of mutant DNA polymerase gamma in patients with mitochondrial DNA depletion. / Taanman, Jan-Willem; Rahman, Shamima; Pagnamenta, Alistair T; Morris, Andrew A M; Bitner-Glindzicz, Maria; Wolf, Nicole I; Leonard, James V; Clayton, Peter T; Schapira, Anthony H V.
In: HUM MUTAT, Vol. 30, No. 2, 2, 2009, p. 248-254.Research output: SCORING: Contribution to journal › SCORING: Journal article › Research › peer-review
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TY - JOUR
T1 - Analysis of mutant DNA polymerase gamma in patients with mitochondrial DNA depletion.
AU - Taanman, Jan-Willem
AU - Rahman, Shamima
AU - Pagnamenta, Alistair T
AU - Morris, Andrew A M
AU - Bitner-Glindzicz, Maria
AU - Wolf, Nicole I
AU - Leonard, James V
AU - Clayton, Peter T
AU - Schapira, Anthony H V
PY - 2009
Y1 - 2009
N2 - We studied six unrelated children with depletion of mitochondrial DNA (mtDNA). They presented with Leigh syndrome, infantile hepatocerebral mtDNA depletion syndrome, or Alpers-Huttenlocher syndrome. Several genes have been implicated in mtDNA depletion. Screening of candidate genes indicated that all six patients were compound heterozygous for missense mutations in the gene for the catalytic subunit of DNA polymerase gamma (POLG). Three of the identified mutations, c.3328C>T (p.H1110Y), c.3401A>G (p.H1134R), and c.3406G>A (p.E1136K), have not been reported earlier. To investigate the functional consequences of the mutations, we carried out a series of biochemical assays in cultured fibroblasts. These studies revealed that fibroblast cultures from the patients with infantile hepatocerebral mtDNA depletion syndrome progressively lost their mtDNA during culturing, whereas fibroblast cultures from patients presenting with Leigh syndrome or Alpers-Huttenlocher syndrome had reduced but stable levels of mtDNA. DNA polymerase gamma activity was below the normal range in all patient cultures, except for one; however, this culture showed low levels of the heterodimeric enzyme and poor DNA polymerase gamma processivity. Parental fibroblast cultures had normal catalytic efficiency of DNA polymerase gamma, consistent with the observation that all carriers are asymptomatic. Thus, we report the first patient with Leigh syndrome caused by POLG mutations. The cell culture experiments established the pathogenicity of the identified POLG mutations and helped to define the molecular mechanisms responsible for mtDNA depletion in the patients' tissues. The assays may facilitate the identification of those patients in whom screening for POLG mutations would be most appropriate.
AB - We studied six unrelated children with depletion of mitochondrial DNA (mtDNA). They presented with Leigh syndrome, infantile hepatocerebral mtDNA depletion syndrome, or Alpers-Huttenlocher syndrome. Several genes have been implicated in mtDNA depletion. Screening of candidate genes indicated that all six patients were compound heterozygous for missense mutations in the gene for the catalytic subunit of DNA polymerase gamma (POLG). Three of the identified mutations, c.3328C>T (p.H1110Y), c.3401A>G (p.H1134R), and c.3406G>A (p.E1136K), have not been reported earlier. To investigate the functional consequences of the mutations, we carried out a series of biochemical assays in cultured fibroblasts. These studies revealed that fibroblast cultures from the patients with infantile hepatocerebral mtDNA depletion syndrome progressively lost their mtDNA during culturing, whereas fibroblast cultures from patients presenting with Leigh syndrome or Alpers-Huttenlocher syndrome had reduced but stable levels of mtDNA. DNA polymerase gamma activity was below the normal range in all patient cultures, except for one; however, this culture showed low levels of the heterodimeric enzyme and poor DNA polymerase gamma processivity. Parental fibroblast cultures had normal catalytic efficiency of DNA polymerase gamma, consistent with the observation that all carriers are asymptomatic. Thus, we report the first patient with Leigh syndrome caused by POLG mutations. The cell culture experiments established the pathogenicity of the identified POLG mutations and helped to define the molecular mechanisms responsible for mtDNA depletion in the patients' tissues. The assays may facilitate the identification of those patients in whom screening for POLG mutations would be most appropriate.
KW - Humans
KW - Male
KW - Female
KW - Adolescent
KW - Infant
KW - Cells, Cultured
KW - Fibroblasts enzymology
KW - Infant, Newborn
KW - DNA Mutational Analysis
KW - DNA-Directed DNA Polymerase genetics
KW - Mutation genetics
KW - DNA, Mitochondrial genetics
KW - Holoenzymes metabolism
KW - Immunoblotting
KW - Humans
KW - Male
KW - Female
KW - Adolescent
KW - Infant
KW - Cells, Cultured
KW - Fibroblasts enzymology
KW - Infant, Newborn
KW - DNA Mutational Analysis
KW - DNA-Directed DNA Polymerase genetics
KW - Mutation genetics
KW - DNA, Mitochondrial genetics
KW - Holoenzymes metabolism
KW - Immunoblotting
M3 - SCORING: Zeitschriftenaufsatz
VL - 30
SP - 248
EP - 254
JO - HUM MUTAT
JF - HUM MUTAT
SN - 1059-7794
IS - 2
M1 - 2
ER -
