Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2015 May 15;24(10):2796-807.
doi: 10.1093/hmg/ddv041. Epub 2015 Feb 4.

Ganglioside GM3 is essential for the structural integrity and function of cochlear hair cells

Misato Yoshikawa  1 , Shinji Go  2 , Shun-ichi Suzuki  2 , Akemi Suzuki  3 , Yukio Katori  4 , Thierry Morlet  5 , Steven M Gottlieb  6 , Michihiro Fujiwara  7 , Katsunori Iwasaki  7 , Kevin A Strauss  8 , Jin-ichi Inokuchi  9
Affiliations

Ganglioside GM3 is essential for the structural integrity and function of cochlear hair cells

Misato Yoshikawa et al. Hum Mol Genet. .

Abstract

GM3 synthase (ST3GAL5) is the first biosynthetic enzyme of a- and b-series gangliosides. Patients with GM3 synthase deficiency suffer severe neurological disability and deafness. Eight children (ages 4.1 ± 2.3 years) homozygous for ST3GAL5 c.694C>T had no detectable GM3 (a-series) or GD3 (b-series) in plasma. Their auditory function was characterized by the absence of middle ear muscle reflexes, distortion product otoacoustic emissions and cochlear microphonics, as well as abnormal auditory brainstem responses and cortical auditory-evoked potentials. In St3gal5(-/-) mice, stereocilia of outer hair cells showed signs of degeneration as early as postnatal Day 3 (P3); thereafter, blebs devoid of actin or tubulin appeared at the region of vestigial kinocilia, suggesting impaired vesicular trafficking. Stereocilia of St3gal5(-/-) inner hair cells were fused by P17, and protein tyrosine phosphatase receptor Q, normally linked to myosin VI at the tapered base of stereocilia, was maldistributed along the cell membrane. B4galnt1(-/-) (GM2 synthase-deficient) mice expressing only GM3 and GD3 gangliosides had normal auditory structure and function. Thus, GM3-dependent membrane microdomains might be essential for the proper organization and maintenance of stereocilia in auditory hair cells.

PubMed Disclaimer

Figures

Figure 1.
Figure 1.
Marked increase of GSLs in cochlea during postnatal maturation period. (A) Schematic presentation of GSL biosynthesis. (B) High performance-thin-layer chromatograms (HPTLC analysis of gangliosides in cochlea of wild-type mice during the postnatal maturation period. Left panel is neutral glycolipids and right panel is acidic glycolipids. Cochlea were isolated and subjected for the purification of GSL analysis as described in Materials and Methods. Each purified fraction was spotted as 2 mg protein per lane. (C) Mass chromatograms of gangliosides prepared from cochlea of the wild mice at P17. The acidic fraction was analyzed by liquid chromatography-ion trap-mass spectrometry with a NH2 column under the conditions described in the Materials and Methods. The MS1 chromatogram was monitored with theoretical values of [M – H]−1 or [M − 2H]−2 for each ganglioside. The structures indicated in the figure is tentative, and based on information with only molecular related ions indicated with black and, in addition, carbohydrate-related fragment ions indicated with red, and requires the consideration of possibility of d20:1 sphingosine. (D) Summary of stage-specific expression of GSLs during postnatal maturation of cochlea based on the information of Figure 1B and C. Comparative development of the cochlea of both humans and mice are also shown.
Figure 2.
Figure 2.
Upper panel from left to right, T2, fluid-attenuated inversion recovery, diffusion-weighted, and 1H-MR spectroscopy at 1.5 T show normal cortical and subcortical structure, but severe hypomyelination and increased water diffusion throughout the corona radiata, as well as elevated choline (Cho) relative to creatine–phosphocreatine (Cr) and N-acetylaspartate 1H signals (single voxel PRESS, TE 144/TR 1500). (Lower panel) EEG of a 2-year-old child homozygous for ST3GAL5 c.694C>T shows a slow, chaotic background, absent posterior rhythm and no discernable change through a behavioral sleep–wake cycle. Beginning at the dashed red line, there is a burst of generalized, frontal predominant, spike-slow wave discharges at 3 Hz in excess of 450 µV (note voltage sensitivity).
Figure 3.
Figure 3.
Identification of GM3 as an essential molecule for hair cell function. (A) HP-TLC of acidic (left) and neutral (right) GSLs from the cochlea of 4-week-old wild-type (WT), B4galnt1−/− and St3gal5−/− mice. The cochlea of B4galnt1−/− mice were mainly expressed GM3 and GD3. Each purified fraction was spotted 2 mg protein per lane. (B) Auditory-evoked brainstem response (ABR) of wild-type, B4galnt1−/− and St3gal5−/− mice at 4 weeks old. (C and D) Comparison of gangliosides expression in hair cells among wild-type, B4galnt1−/− and St3gal5−/− mice aged 4 weeks. Whole-mount immunostaining was performed. GM1 (C, green) and GM3 (D, green) of IHC (a–c) and OHC (d–f) stereocilia in wild-type (a and d), B4galnt1−/− (b and e) and St3gal5−/− (c and f) mice. In IHC, both GM1 and GM3 were expressed along the stereocilium from the taper region to top (Ca and Da). On the other hand, only GM3 was highly enriched in OHC stereocilia (Dd) without staining of GM1 (Cd). The stereocilia of IHC and OHC in B4galnt1−/− mice were expressed GM3 but not GM1 (Cb and Db). In St3gal5−/− mice, fused stereocilia was observed in IHC (Cc and Dc) and irregular phalloidin staining was observed in OHC (Cf and Df). Notably, the expression levels of GM1 but not GM3 was especially high on the surface of supporting cells (Deiters cell) (Cd and Dd). (E) Confocal images of cross sections of the cochleae in wild-type mice stained for GM3 (green, left panel), GM1 (green, right panel) and propidium iodide to label cell nuclei. (Arrow) Deiters cell, (arrow head) pillar cell. The red color of phalloidin staining (C and D) and propidium iodide (E) has been converted to magenta using Adobe Photoshop CC2014 software.
Figure 4.
Figure 4.
Degeneration of OHC in St3gal5 null mice. (A) Confocal images of the basal coil of phalloidin-stained cochlear whole mounts from St3gal5+/+ and −/− mice at P1 and P3. Defects in structure of hair bundle are first in the outer hair cells (arrows) at P3. (B) SEM images of OHC of St3gal5+/+ and St3gal5−/− mice at P8 and 17. At P8, the blebs at the vestigial kinocilium location were observed (arrows), but disappeared at P17. (C) Expression of acetylated tubulin. Confocal images from showing stereocilia of St3gal5+/+ and −/− mice stained for acetylated tubulin (green) and F-actin (phalloidin, magenta) at P3 and P8. At P3, hair cells of St3gal5+/+ and −/− mice were expressed kinocilia with or without degeneration of stereocilia. At P8, kinocilia were disappeared from OHC in both St3gal5+/+ and −/− mice. Three rows of OHCs are indicated by the white arrowheads. (D) TUNEL staining. Apoptotic cell was not detected in both St3gal5+/+ and −/− mice at P8. (E) TEM images of OHC in St3gal5+/+ and −/− mice at P17. The red color of phalloidin staining (C) and propidium iodide (D) has been converted to magenta using Adobe Photoshop CC2014 software.
Figure 5.
Figure 5.
Fused stereocilia formation of IHC in St3gal5 null mice. (A)The stereocilia of the IHCs of St3gal5−/− mice were fused after P12. Arrows indicate the fused stereocilia. (B) TEM images of the IHC stereocilia in St3gal5+/+ and −/− mice at P17.
Figure 6.
Figure 6.
Dislocalization of PTPRQ and myosinVI in the stereocilia of St3gal5 null mice. Confocal images showing stereocilia of IHCs and OHCs of St3gal5+/+ and −/− mice stained for myosinVI (A) PTPRQ (B) (green) and F-actin (phalloidin, magenta). (C) Schematic images for dislocalization of PTPRQ and myosin VI in the IHC stereocilia of St3gal5−/− mouse. The red color of phalloidin staining (A and B) has been converted to magenta using Adobe Photoshop CC2014 software.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Roberts W.M., Howard J., Hudspeth A.J. (1988) Hair cells: transduction, tuning, and transmission in the inner ear. Annu. Rev. Cell Biol., 4, 63–92. - PubMed
    1. Hudspeth A.J. (1997) How hearing happens. Neuron, 19, 947–950. - PubMed
    1. Hakomori S. (1981) Glycosphingolipids in cellular interaction, differentiation, and oncogenesis. Annu. Rev. Biochem., 50, 733–764. - PubMed
    1. Schengrund C.L. (1990) The role(s) of gangliosides in neural differentiation and repair: a perspective. Brain. Res. Bull., 24, 131–141. - PubMed
    1. Kawai H., Allende M.L., Wada R., Kono M., Sango K., Deng C., Miyakawa T., Crawley J.N., Werth N., Bierfreund U., Sandhoff K., Proia R.L. (2001) Mice expressing only monosialoganglioside GM3 exhibit lethal audiogenic seizures. J. Biol. Chem., 276, 6885–6888. - PubMed

Publication types

MeSH terms

Substances

Supplementary concepts