Targeted knockout of the mouse beta B2-crystallin gene (Crybb2) induces age-related cataract
Jun Zhang 1,
Jing Li 2,
Caiguo Huang 2,
Lian Xue 1,
Yajun Peng 3,
Qing Fu 3,
Li Gao 4,
Jianrong Zhang 1,
and
wenjie li 5*
1 Central Laboratory, Shanghai, China
2 Department of Biochemistry and Molecular Biology, Shanghai, China
3 Department of Ophthalmology, Shanghai, China
4 Department of Pathology, Shanghai, China
5 Central Laboratory, 174 Changhai Road, Central Laboratory, Changhai Hospital, Shanghai 200433, People?s Republic of China, Shanghai, 200433, China
* To whom correspondence should be addressed. E-mail: schoman123{at}yahoo.com.cn.
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Abstract |
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PURPOSE. The beta B2-crystallin gene (Crybb2) is expressed at an increasing level in the postnatal lens cortex. It has been assumed to function as structural protein, although this has not been directly tested. Here we begin to examine the in vivo functions of beta B2-crystallin by generating mice with a targeted disruption of Crybb2. METHODS. Gene targeting in embryonic stem cells was used to generate mouse lines in which Crybb2 was deleted. Gene structure and protein expression were analyzed by PCR, immunoblot, and two-dimensional gel electrophoresis. Knockout mice were screened for cataract with slit lamp biomicroscopy. Microstructure of lens was analyzed by scanning and transmission electron microscopy. The resistance of crystallins in knockout mice to heat-induced denaturation and oxidative stress was examined. RESULTS. The lens appeared to develop normally in the first months of life. In older animals, the weight and axial diameter of the lenses of knockout mice were significantly smaller than in wild type. Cataracts were formed in the posterior and anterior cortex several months after birth and cataract severity increase with age. The thermal stability of the supernatant of a lens homogenate was mildly compromised. The knockout lenses also showed decreased resistance to oxidative stress. CONCLUSIONS. beta B2 crystallin is not essential for the normal development of a transparent lens in the mouse. It plays an increasingly important role in maintaining the transparency of lens after birth, possibly by the interaction with other crystallins to increase their resistance to thermal denaturation and oxidative stress.
Key Words:
lens proteins, cataractogenesis, knockout animals, beta B2-crystallin
