Intracellular distribution of transgenic bacterial beta-galactosidase in central nervous system neurons and neuroglia.
Bacterial beta-galactosidase is widely used as a marker for gene expression and in cell tracing experiments. In a survey of three transgenic mouse lines expressing beta-galactosidase in the central nervous system (CNS) under the control of different promoters, we find substantial variation in the intracellular distribution of the lacZ protein. In line M beta P5, transgene beta-galactosidase expression is driven by a promoter/enhancer fragment from the oligodendrocyte-specific myelin basic protein gene; however, electron microscopy of histochemically stained preparations reveals transgene expression not only in oligodendrocytes but also in some neurons. Immunofluorescence and immunoperoxidase staining show the beta-galactosidase protein distributed throughout the perikaryal cytoplasm of oligodendrocytes and in processes reaching to myelin sheaths. By contrast, immunoreactive protein appears restricted in neurons to one or a few small perikaryal immunoreactive granules. The granules are visible in the electron microscope as amorphous inclusion bodies of moderate electron density and lack a limiting membrane. Histochemical staining patterns with X-gal and Bluo-gal echoed the protein distribution: diffuse distribution of enzyme protein yielded cells filled with substrate, while punctate enzyme distribution yielded restricted or punctate histochemical staining. Examination of two other lines using different promoter/enhancers to drive expression in the CNS showed both diffuse and punctate beta-galactosidase immunolocalization and histochemical staining. The amount of protein synthesized or other properties, yet unidentified, intrinsic to the target cells may determine the intracellular distribution of beta-galactosidase. In retroviral marking studies, clone members have been identified as those cells filled with X-gal reaction product. This approach may underestimate both clone size and the minimum number of divisions separating the members of each clone.
Friedrich
V L
VL
Brookdale Center for Molecular Biology, Mount Sinai School of Medicine, New York, New York 10029-6574.
Holstein
G R
GR
Li
X
X
Gow
A
A
Kelley
K A
KA
Lazzarini
R A
RA
D000818:Animals; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics; D002460:Cell Line; D002490:Central Nervous System / Q000166:cytology* / Q000254:growth & development / Q000378:metabolism; D005786:Gene Expression Regulation; D005813:Genes, Synthetic; D051379:Mice; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D008822:Mice, Transgenic; D009419:Nerve Tissue Proteins / Q000096:biosynthesis* / Q000235:genetics; D009457:Neuroglia / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis*; D013234:Stem Cells / Q000378:metabolism; D013347:Subcellular Fractions / Q000737:chemistry; D001616:beta-Galactosidase / Q000096:biosynthesis* / Q000235:genetics
D001426:Bacterial Proteins; D009419:Nerve Tissue Proteins; D011993:Recombinant Fusion Proteins; D001616:beta-Galactosidase