Limits...
Cystathionine beta-synthase p.S466L mutation causes hyperhomocysteinemia in mice.

Gupta S, Wang L, Hua X, Krijt J, Kozich V, Kruger WD - Hum. Mutat. (2008)

Bottom Line: Surprisingly, CBS mRNA was significantly elevated in Tg-S466L animals compared to Tg-hCBS, implying that the reduction in p.S466L protein was occurring due to posttranscriptional mechanisms.In Tg-S466L animals with detectable liver CBS, the enzyme formed tetramers and was active, but lacked inducibility by S-adenosylmethionine (AdoMet).However, even in Tg-S466L animals that had in vitro liver CBS activity equivalent to Tg-hCBS animals there was significant elevation of serum tHcy.

View Article: PubMed Central - PubMed

Affiliation: Division of Population Science, Fox Chase Cancer Center, Philadelphia, Pennsylvania 19111, USA.

ABSTRACT
Missense mutations in the cystathionine beta-synthase (CBS) gene are the most common cause of clinical homocystinuria in humans. The p.S466L mutation was identified in a homocystinuric patient, but enzymatic studies with recombinant protein show this mutant to be highly active. To understand how this mutation causes disease in vivo, we have created mice lacking endogenous mouse CBS and expressing either wild-type (Tg-hCBS) or p.S466L (Tg-S466L) human CBS under control of zinc inducible metallothionein promoter. In the presence of zinc, we found that the mean serum total homocysteine (tHcy) of Tg-S466L mice was 142+/-55 microM compared to 16+/-13 microM for hCBS mice. Tg-S466L mice also had significantly higher levels of total free homocysteine and S-adenosylhomocysteine in liver and kidney. Only 48% of Tg-S466L mice had detectable CBS protein in the liver, whereas all the Tg-hCBS animals had detectable protein. Surprisingly, CBS mRNA was significantly elevated in Tg-S466L animals compared to Tg-hCBS, implying that the reduction in p.S466L protein was occurring due to posttranscriptional mechanisms. In Tg-S466L animals with detectable liver CBS, the enzyme formed tetramers and was active, but lacked inducibility by S-adenosylmethionine (AdoMet). However, even in Tg-S466L animals that had in vitro liver CBS activity equivalent to Tg-hCBS animals there was significant elevation of serum tHcy. Our results show that p.S466L causes homocystinuria by affecting both the steady state level of CBS protein and by reducing the efficiency of the enzyme in vivo.

Show MeSH

Related in: MedlinePlus

Correlation between CBS activity, tetramer formation, and serum tHcy. A: Correlation between CBS tetramer density and CBS activity (in presence of AdoMet) in the livers of mice of respective genotype. Black dots represent Tg-hCBS mice and white dots represent Tg-S466L mice. B: Correlation between liver CBS activity (in presence of AdoMet) and serum tHcy in Tg-hCBS (n = 8) and Tg-S466L (n = 9) mice. CBS activity has been measured in the terms of micromoles (µmol) of cystathionine formed per milligram of protein per hr.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2630375&req=5

fig05: Correlation between CBS activity, tetramer formation, and serum tHcy. A: Correlation between CBS tetramer density and CBS activity (in presence of AdoMet) in the livers of mice of respective genotype. Black dots represent Tg-hCBS mice and white dots represent Tg-S466L mice. B: Correlation between liver CBS activity (in presence of AdoMet) and serum tHcy in Tg-hCBS (n = 8) and Tg-S466L (n = 9) mice. CBS activity has been measured in the terms of micromoles (µmol) of cystathionine formed per milligram of protein per hr.

Mentions: We examined the relationship between the amount of CBS tetramer present in liver homogenates from Tg-S466L and Tg-hCBS animals and CBS activity. We found that there was a strong linear relationship between the level of tetramer and CBS activity for both Tg-hCBS and Tg-S466L animals (R2 = 0.96; Fig. 5A). Both the wild-type and the p.S466L animals appear to lie along the same line. From these findings we conclude that the amount of tetramer formed is a good predictor of CBS activity.


Cystathionine beta-synthase p.S466L mutation causes hyperhomocysteinemia in mice.

Gupta S, Wang L, Hua X, Krijt J, Kozich V, Kruger WD - Hum. Mutat. (2008)

Correlation between CBS activity, tetramer formation, and serum tHcy. A: Correlation between CBS tetramer density and CBS activity (in presence of AdoMet) in the livers of mice of respective genotype. Black dots represent Tg-hCBS mice and white dots represent Tg-S466L mice. B: Correlation between liver CBS activity (in presence of AdoMet) and serum tHcy in Tg-hCBS (n = 8) and Tg-S466L (n = 9) mice. CBS activity has been measured in the terms of micromoles (µmol) of cystathionine formed per milligram of protein per hr.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2630375&req=5

fig05: Correlation between CBS activity, tetramer formation, and serum tHcy. A: Correlation between CBS tetramer density and CBS activity (in presence of AdoMet) in the livers of mice of respective genotype. Black dots represent Tg-hCBS mice and white dots represent Tg-S466L mice. B: Correlation between liver CBS activity (in presence of AdoMet) and serum tHcy in Tg-hCBS (n = 8) and Tg-S466L (n = 9) mice. CBS activity has been measured in the terms of micromoles (µmol) of cystathionine formed per milligram of protein per hr.
Mentions: We examined the relationship between the amount of CBS tetramer present in liver homogenates from Tg-S466L and Tg-hCBS animals and CBS activity. We found that there was a strong linear relationship between the level of tetramer and CBS activity for both Tg-hCBS and Tg-S466L animals (R2 = 0.96; Fig. 5A). Both the wild-type and the p.S466L animals appear to lie along the same line. From these findings we conclude that the amount of tetramer formed is a good predictor of CBS activity.

Bottom Line: Surprisingly, CBS mRNA was significantly elevated in Tg-S466L animals compared to Tg-hCBS, implying that the reduction in p.S466L protein was occurring due to posttranscriptional mechanisms.In Tg-S466L animals with detectable liver CBS, the enzyme formed tetramers and was active, but lacked inducibility by S-adenosylmethionine (AdoMet).However, even in Tg-S466L animals that had in vitro liver CBS activity equivalent to Tg-hCBS animals there was significant elevation of serum tHcy.

View Article: PubMed Central - PubMed

Affiliation: Division of Population Science, Fox Chase Cancer Center, Philadelphia, Pennsylvania 19111, USA.

ABSTRACT
Missense mutations in the cystathionine beta-synthase (CBS) gene are the most common cause of clinical homocystinuria in humans. The p.S466L mutation was identified in a homocystinuric patient, but enzymatic studies with recombinant protein show this mutant to be highly active. To understand how this mutation causes disease in vivo, we have created mice lacking endogenous mouse CBS and expressing either wild-type (Tg-hCBS) or p.S466L (Tg-S466L) human CBS under control of zinc inducible metallothionein promoter. In the presence of zinc, we found that the mean serum total homocysteine (tHcy) of Tg-S466L mice was 142+/-55 microM compared to 16+/-13 microM for hCBS mice. Tg-S466L mice also had significantly higher levels of total free homocysteine and S-adenosylhomocysteine in liver and kidney. Only 48% of Tg-S466L mice had detectable CBS protein in the liver, whereas all the Tg-hCBS animals had detectable protein. Surprisingly, CBS mRNA was significantly elevated in Tg-S466L animals compared to Tg-hCBS, implying that the reduction in p.S466L protein was occurring due to posttranscriptional mechanisms. In Tg-S466L animals with detectable liver CBS, the enzyme formed tetramers and was active, but lacked inducibility by S-adenosylmethionine (AdoMet). However, even in Tg-S466L animals that had in vitro liver CBS activity equivalent to Tg-hCBS animals there was significant elevation of serum tHcy. Our results show that p.S466L causes homocystinuria by affecting both the steady state level of CBS protein and by reducing the efficiency of the enzyme in vivo.

Show MeSH
Related in: MedlinePlus