Gap junction structures. IV. Asymmetric features revealed by low-irradiation microscopy.

Abstract

Micrographs of mouse liver gap junctions, isolated with detergents, and negatively stained with uranyl acetate, were recorded by low-irradiation methods. Fourier-averaged micrographs of the hexagonal junction lattice show skewed, hexameric connexons with less stain at the 3-fold axis than at the 6 indentations between the lobes of the connexon image. These substructural features, not clearly observed previously, are acutely sensitive to irradiation. After an electron dose less than that normally used in microscopy, the image is converted to the familiar doughnut shape, with a darkly stained center and a smooth hexagonal outline, oriented with mirror symmetry in the lattice. Differences in appearance among 25 reconstructed images from low-irradiation micrographs illustrate variation in staining of the connexon channel and the space between connexons. Consistently observed stain concentration at 6 symmetrically related sites .apprx. 34 .ANG. from the connexon center, 8.degree. to the right or left of the (1, 1) lattice vector, may reveal an intrinsic asymmetric feature of the junction structure. The unexpected skewing of the 6-lobed connexon image suggests that the pair of hexagonal membrane arrays that form the junction may not be structurally identical. Because the projected image of the connexon pair itself appears mirror-symmetric, each pair may consist of 2 identical connexon hexamers related by local (noncrystallographic) 2-fold axes in the junctional plane at the middle of the gap. All connexons may be chemically identical, but their packing in the hexagonal arrays on the 2 sides of the junction appears to be nonequivalent.