Skip to main content
NCBI home page
microPublication Biology logoLink to microPublication Biology
. 2022 Nov 12;2022:10.17912/micropub.biology.000678. doi: 10.17912/micropub.biology.000678

Tubulin posttranslational modifications modify the atypical spermatozoon centriole

Katerina A Turner 1, Derek F Kluczynski 1, Ryan J Hefner 1, Rami B Moussa 1, Julia N Slogar 1, Joice B Thekkethottiyil 1, Haley D Prine 1, Emily R Crossley 1, Lucas J Flanagan 1, Marlena M LaBoy 1, Mia B Moran 1, Taylor G Boyd 1, Benjamin A Kujawski 1, Kelsie Ruble 1, John M Pap 1, Ankit Jaiswal 1, Tariq A Shah 1, Puneet Sindhwani 1, Tomer Avidor-Reiss 1,§
Reviewed by: Anonymous
PMCID: PMC9700210  PMID: 36444375

Abstract

Sperm cells are transcriptionally and translationally silent. Therefore, they may use one of the remaining mechanisms to respond to stimuli in their environment, the post-translational modification of their proteins. Here we examined three post-translational modifications, acetylation, glutamylation, and glycylation of the protein tubulin in human and cattle sperm. Tubulin is the monomer that makes up microtubules, and microtubules constitute the core component of both the sperm centrioles and the axoneme. We found that the sperm of both species were labeled by antibodies against acetylated tubulin and glutamylated tubulin.

Figure 1. Acetylated tubulin and glutamylated tubulin are observed in both cattle and human sperm centrioles, while glycylated tubulin staining is variable .

Figure 1. Acetylated tubulin and glutamylated tubulin are observed in both cattle and human sperm centrioles, while glycylated tubulin staining is variable

Open in a new tab

A ) The three main parts of cattle and human sperm are the head (H), neck (N), and tail (T). The head contains the nucleus (Nu), the proximal centriole (PC) and distal centriole (DC) are within the neck, and the tail consists of the axoneme (Ax), which extends through the midpiece (M), the principal piece (P), and end piece (E). B ) The proximal centriole is barrel-shaped, comprised of nine microtubule triplets, and is nearly perpendicular to the anterior side of the distal centriole. The distal centriole has an atypical structure with two protein rods (Ro), has nine splayed doublet microtubules, and is directly connected to the axoneme. C-H ) Immunofluorescence staining was used to observe different proteins and posttranslational modifications in cattle ( C , E , G ) and human ( D , F , H ) sperm. POC1B (magenta) is observed almost exclusively in the proximal and distal centrioles. Tubulin (red) labels the PC, DC, and axoneme. C-D ) The antibody (Thermo Fisher Scientific, MA533079) against acetylated tubulin (Ace Tub, green) was primarily detected in the PC, DC, and especially in the axoneme of cattle ( C ) and human ( D ) sperm. E-F ) The antibody (Adipogen, AG-20B-0020-C100) against glutamylated tubulin (Glu Tub, green) labeled the PC and DC of cattle ( E ) and human ( F ) sperm. G ) The antibody (EMD Millipore, MABS277) against glycylated tubulin (Gly Tub, green) was not detected in any centriolar components of cattle. H ) The antibody (Adipogen, rabbit ab, AG-25B-0034-C100) against monoglycylated tubulin (Gly Tub, green) was not detected in the PC of human sperm, but there is some signal in the midpiece and DC. Note that only the tail structures labeled by the anti-posttranslational modification antibody are annotated in the low magnification panels.

Description

Infertility affects 12-15% of couples; furthermore, one-third of couples have unexplained infertility (Thoma et al. 2013; Pandruvada et al. 2021). One potential explanation for unexplained infertility is anomalies in the sperm centrioles. Sperm centrioles are essential for sperm movement and behavior during swimming and microtubule organization in the early embryo development (Terada et al. 2004; Chemes and Alvarez Sedo 2012; Cavazza et al. 2021).

It was previously thought that there was only one centriole, the proximal centriole (PC), in mammalian sperm because it was the only recognizable centriole with the canonical barrel shape. Recently it was found that human and cattle sperm have a second, atypical, fan-shaped centriole, the distal centriole (DC) (Fishman et al. 2018) ( Fig 1A-B ). The distal centriole and proximal centriole form a dynamic basal complex within the sperm neck that mediates the tail beating, generating a head twitching behavior (Khanal et al. 2021). In this process, the distal centriole’s left-sided microtubules move up and down relative to the distal centriole’s right-sided microtubules, pushing the rest of the dynamic basal complex. However, the precise contribution of the distal centriole and dynamic basal complex to male infertility is unknown.

Identifying the contribution of sperm centrioles to male infertility depends on identifying appropriate markers and quantifying their relative abundance in the sperm centrioles (Turner et al. 2021). These markers can be structural proteins or their posttranslational modifications (Jaiswal et al. 2022). Microtubules, composed of the protein tubulin, make up the main structure of the centriole. Post-translational modifications of tubulin can change the overall properties of the microtubule (Janke and Magiera 2020). Microtubules undergo mechanical stress as part of sperm swimming, the distal centriole microtubules, in particular, are under mechanical stress as part of the dynamic basal complex (Khanal et al. 2021). Given that tubulin modifications can affect microtubule properties, we wondered if these modifications are present in sperm distal centriole.

Interestingly, the enzymes responsible for the tubulin modifications acetylation, glutamylation, and glycylation are present in the sperm, particularly near the centrioles, suggesting the modifications may be occurring. Acetylation is mediated by acetyltransferase and αTAT1 (Kalebic et al. 2013), which are present in sperm (Chawan et al. 2020). Recently, it was shown that protein acetylation protects sperm from spontaneous acrosome reactions (Bowker et al. 2022). Glutamylation is mediated by glutamylases (TTLL 1, 4, 5, 6, 7, 9, and 11) localized at basal bodies, although TTLL 4, 5, 6, and 7 are also localized to the cilia (van Dijk et al. 2007) TTLL1 is found in the sperm and is essential for sperm flagella function (Vogel et al. 2010). Tubulin glycylation is mediated by TTLL3 and TTLL8 (Wloga et al. 2009; Gadadhar et al. 2017). Mouse knockouts of both TTLL3 and TTLL8 have abnormal sperm axonemal dynein activity, flagellar beat, and male fertility (Gadadhar et al. 2021). Therefore, tubulin posttranslational modifications may be present in the sperm distal centriole and useful for developing diagnostics and treatments for centriole-based infertility.

Here, we investigated the presence of these modifications in cattle and human sperm by studying their colocalization with known centriolar biomarkers, tubulin and POC1B. As expected, the anti-tubulin and anti-POC1B antibodies labeled two spots in the sperm neck, the distal centriole and proximal centriole, of cattle and humans (Khanal et al. 2021) ( Fig. 1 ). POC1B immunoreactivity is enriched in the distal centriole relative to the proximal centriole (the ratio of distal centriole: proximal centriole, 4.77 ± 2.61, N=101 bovine sperm; 1.24 ± 0.41, N=198 human sperm), this enrichment is more pronounced in bovine than in human. Similarly, Tubulin immunoreactivity is enriched in the proximal centriole relative to the axoneme more pronouncedly in bovine than in human (the ratio of proximal centriole:(proximal centriole + axoneme), 0.91 ± 0.12, N=101 bovine sperm; 0.62 +/- 0.17, N=198 human sperm).

Tubulin is acetylated at lysine residue Lys 40 of the a-tubulin subunit (LeDizet and Piperno 1987), which is in the microtubule lumen (Nogales et al. 1999; Soppina et al. 2012). Acetylation stabilizes microtubules by providing flexibility and resistance to mechanical stress (Portran et al. 2017; Xu et al. 2017; Eshun-Wilson et al. 2019). Tubulin acetylation is reported in centrioles of somatic cells (Sullenberger et al. 2020) and sperm (Fishman et al. 2018). However, most studies used one antibody that recognizes tubulin acetylation – mouse monoclonal antibody 6-11B-1 (Piperno et al. 1987). Here, we tested a second antibody – rabbit monoclonal antibody (Thermo Fisher Scientific, MA533079). Similar to previous studies with antibody 6-11B-1 in sperm, we found that MA533079 labeled the neck and throughout the tail of cattle and human sperm ( Fig. 1C-D ). Also, like in previous studies with antibody 6-11B-1, we found that in the neck, MA533079 labeled two spots, the sperm centrioles, colocalizing with POC1B in cattle (98% of cells; N=272; 4 independent stainings) and human sperm (100% of cells, N= 96; 3 independent stainings). Tubulin acetylation staining is weaker in the midpiece axoneme compared to the centrioles in cattle (the ratio of proximal centriole:(proximal centriole + axoneme), 0.58±0.18, N=119 sperm) and human sperm (the ratio of proximal centriole:(proximal centriole + axoneme), 0.54±0.19, N=96 sperm; 3 independent stainings).

Tubulin is glutamylated in the C’ terminus at glutamate residue Glu 445 of the a-tubulin subunit, which is on the microtubule exterior (Edde et al. 1990). Tubulin glutamylation stabilizes microtubules (Tremoleda et al. 2003; Hamel et al. 2017). Tubulin glutamylation is present in somatic cell centrioles (Bobinnec et al. 1998) and the sperm proximal centriole (Fouquet et al. 1994); however, its status in sperm distal centriole remains unknown. We found that anti-glutamylated tubulin antibody GT335 labeled the sperm neck consistently and sometimes the tail midpiece or endpiece in humans and consistently in the endpiece in cattle ( Fig. 1E-F ). In the neck, anti-glutamylated Tubulin antibody GT335 labeled two spots, colocalizing with POC1B in cattle (81.5% of 146 cells, 4 independent stainings) and human sperm (73% of N=90 sperm total from 3 men). This staining is enriched in the proximal centriole compared to the midpiece axoneme in cattle (the ratio of proximal centriole:(proximal centriole + axoneme), 0.85 ± 0.21, N=136 sperm) and in human (the ratio of proximal centriole:(proximal centriole + axoneme), 0.56 ± 0.18, N=90 sperm).

Tubulin glycylation is found at glutamate residues Glu 437 , Glu 438 , Glu 439 , and Glu 441 in the β-tubulin subunit and glutamate residues Glu 445 , Glu 446 , and Glu 448 in the α-tubulin (Redeker et al. 1994) which are all found in the microtubule exterior (Magiera and Janke 2014). The specific effect of tubulin glycylation on microtubules is unclear, but it is strongly suggested that glycylation regulates cilia and flagella length and assembly (Thazhath et al. 2004; Gadadhar et al. 2021). Tubulin glycylation has not been found in somatic cell centrioles (Gadadhar et al. 2017). Glycylation affects sperm flagellar beat (Gadadhar et al. 2021), but it was not observed in human sperm proximal centriole using antibody AXO49 mAb and TAP952 mAb, although its presence in the distal centriole remains unknown (Kann et al. 1998). We tested two distinct antibodies that label glycylation and found inconsistent labeling. We found that mouse anti-glycylated Tubulin antibody (EMD Millipore, MABS277) diffusely labeled the neck and the tail at the mid-piece and end-piece of cattle sperm (100%, N=19, 3 independent with increasing antibody concentrations) ( Fig. 1G ). In human sperm, it did not label the sperm at all (0%, N=10, 3 independent stainings). In contrast, the rabbit anti-glycylated tubulin antibody Adipogen, AG-25B-0034-C100 labeled the cattle sperm neck, midpiece and end piece (94%, N=102, 2 independent experiments). In humans the anti-rabbit glycylated tubulin antibody Adipogen, AG-25B-0034-C100 labeled the midpiece and distal centriole ( Fig. 1H ). Midpiece labeling was observed in 100% of cells, and distal centriole labeling was observed in 93.3% of cells (N=15). Overall glycylation in the atypical centriole varies depending on species and antibody and therefore is questionable.

Altogether, our data suggests that acetylation and glutamylation are present in the atypical distal centriole and can be used as sperm centriolar biomarkers in a species-specific manner.

Methods

Immunofluorescence and quantification was completed as previously described (Turner et al. 2021; Jaiswal et al. 2022), and the antibody dilutions are described in the reagents section.

Reagents

Primary Antibodies:

Anti-POC1B made in Mouse (Thermo Fisher Scientific, H00282809-B01P) (Diluted 1 to 300 in cattle and 1 to 100-300 in human sperm). This mouse polyclonal antibody was raised against a full-length human POC1B (aka WDR51B). The antibody specificity was demonstrated by the absence of immunoreactivity in the sperm centriole of rabbits with a POC1B mutation but was immunoreactive in the sperm centriole of control rabbits in our lab.

Anti-POC1B made in Rabbit (Thermo Fisher Scientific, PA5-24495) (Diluted 1 to 100 in cattle and human sperm). The specificity of the antibody was demonstrated in multiples studies showing specific labeling of the centriole (Chang et al. 2016; Liu et al. 2020; Amargant et al. 2021).

Anti-Tubulin made in Sheep (Cytoskeleton, Inc., ATN02) (Diluted 1 to 600 in cattle and 1 to 300 up to 600 in human sperm). The specificity of the antibody toward microtubules was demonstrated in reference (Piroli et al. 2014; Lobert et al. 2022).

Anti-Acetylated Tubulin made in Rabbit (Thermo Fisher Scientific, MA5-33079) (Diluted 1 to 100 in cattle and human sperm). The specificity of the antibody was demonstrated by labeling purified Chlamydomoanas reinhardtii centrioles (Mahecic et al. 2020).

Anti-Polyglutamylated Tubulin made in Mouse (Adipogen, AG-20B-0020-C100) (Diluted 1 to 50 up to 100 in cattle and 1 to 100 in human sperm). The specificity of the antibody was demonstrated by attenuating the activity of TTLL4, the enzyme mediating microtubule polyglutamylation (Arnold et al. 2020).

Anti-Glycylated Tubulin made in Rabbit (Adipogen, AG-25B-0034-C100, aka Gly-pep1) (Diluted 1 to 25 up to 100 in cattle and 1 to 100 in human sperm). The specificity of the antibody was demonstrated by attenuating the activity of TTLL3 and TTLL8 the enzymes mediating microtubule monoglycylation (Gadadhar et al. 2017).

Anti-Monoglycylated Tubulin made in Mouse (EMD Millipore, MABS277, aka TAP952) (Diluted 1 to 25 up to 100 in cattle and 1 to 100 in human sperm). The specificity of the antibody was demonstrated by attenuating the activity of TTLL3 the enzyme mediating microtubule monoglycylation (Wloga et al. 2009; Gadadhar et al. 2017).

Secondary Antibodies:

Anti-Sheep Alexa 555 made in Donkey (Thermo Fisher Scientific, A-21436) (Diluted 1 to 1000)

Anti-Mouse DyLight 488 made in Donkey (Thermo Fisher Scientific, SA5-10166) (Diluted 1 to 400)

Anti-Rabbit DyLight 650 made in Donkey (Thermo Fisher Scientific, SA5-10041) (Diluted 1 to 400)

Solutions:

Washing Solution: PBS

Permeabilization Solution: PBS with 0.3% Triton X-100 (PBST) (Sigma Aldrich, 9002-93-1)

Blocking Solution: PBST with 1% BSA (PBSTb) (CHEM-IMPEX INT’L, 00535)

Sperm: Frozen human and cattle sperm were stored in liquid nitrogen and obtained from UToledo andrology clinic and Select Sires Inc. IRB number 202366-UT and 300220-UT, and IBC number108074-UT

Cattle sperm was removed from the straws and human sperm was removed from cryovials. Both types of samples were put onto slides in 5 steps: 1. Straw or cryovials were placed in 37°C water; 2. Sperm was washed using a 40/80 density gradient (Nidacon, Cat # PS40-100 and PS80-100); 3. The pellet was collected and washed in sperm washing media (Nidacon, Cat # PSW-100), then resuspended in M199 (Sigma-Aldrich, Cat # M7528-500ML); 5. Sperm was added to the slide and then dropped into liquid nitrogen where it was stored until use.

Fixation: Methanol (-20°C) (Fisher Chemical, A412P-4)

A humidity chamber is a plastic container that is layered with wet paper towels covered with a lid

Material:

Confocal Microscope: Leica SP8, Sperm images were taken at a magnification of 640× and zoom of 6×, with 512 × 512-pixel density, 3x zoom and 1024x1024-pixel density or 0.75x zoom and 4096x4096-pixel density

Using Photoshop, immunofluorescence sperm panels were cropped to:

- Low magnification: 525 x 1050 pixels and adjusted to 1-inch x 2 inches

- Medium magnification: 150 x 300 pixels and adjusted to 1-inch x 2 inches

- High magnification: 75 x 75 pixels and adjusted to 1-inch x 1 inch.

Adobe Illustrator was used to create sperm images.

Parafilm Wax (VWR, 52858-032)

Cover Slip (VWR, 48366-205)

Slides (Home Science Tools, MS-SLFRO72)

Nail Polish (EMS Diasum, 72180)

Mounting Media made of Fluoroshield with DAPI (Sigma-Aldrich, F6057-20ML)

Hoechst (Thermo Fisher Scientific, H1399) (Diluted 1 to 2000)

LAS X Software Leica using photon counting and BrightR.

Statistical methods: Experiments were completed 2-4 times; we calculated averages and standard deviations using Microsoft Excel.

Primary Antibodies:

Anti-POC1B made in Mouse (Thermo Fisher Scientific, H00282809-B01P) (Diluted 1 to 300 in cattle and 1 to 100-300 in human sperm). This mouse polyclonal antibody was raised against a full-length human POC1B (aka WDR51B). The antibody specificity was demonstrated by the absence of immunoreactivity in the sperm centriole of rabbits with a POC1B mutation but was immunoreactive in the sperm centriole of control rabbits in our lab.

Anti-POC1B made in Rabbit (Thermo Fisher Scientific, PA5-24495) (Diluted 1 to 100 in cattle and human sperm). The specificity of the antibody was demonstrated in multiples studies showing specific labeling of the centriole (Chang et al. 2016; Liu et al. 2020; Amargant et al. 2021).

Anti-Tubulin made in Sheep (Cytoskeleton, Inc., ATN02) (Diluted 1 to 600 in cattle and 1 to 300 up to 600 in human sperm). The specificity of the antibody toward microtubules was demonstrated in reference (Piroli et al. 2014; Lobert et al. 2022).

Anti-Acetylated Tubulin made in Rabbit (Thermo Fisher Scientific, MA5-33079) (Diluted 1 to 100 in cattle and human sperm). The specificity of the antibody was demonstrated by labeling purified Chlamydomoanas reinhardtii centrioles (Mahecic et al. 2020).

Anti-Polyglutamylated Tubulin made in Mouse (Adipogen, AG-20B-0020-C100) (Diluted 1 to 50 up to 100 in cattle and 1 to 100 in human sperm). The specificity of the antibody was demonstrated by attenuating the activity of TTLL4, the enzyme mediating microtubule polyglutamylation (Arnold et al. 2020).

Anti-Glycylated Tubulin made in Rabbit (Adipogen, AG-25B-0034-C100, aka Gly-pep1) (Diluted 1 to 25 up to 100 in cattle and 1 to 100 in human sperm). The specificity of the antibody was demonstrated by attenuating the activity of TTLL3 and TTLL8 the enzymes mediating microtubule monoglycylation (Gadadhar et al. 2017).

Anti-Monoglycylated Tubulin made in Mouse (EMD Millipore, MABS277, aka TAP952) (Diluted 1 to 25 up to 100 in cattle and 1 to 100 in human sperm). The specificity of the antibody was demonstrated by attenuating the activity of TTLL3 the enzyme mediating microtubule monoglycylation (Wloga et al. 2009; Gadadhar et al. 2017).

Secondary Antibodies:

Anti-Sheep Alexa 555 made in Donkey (Thermo Fisher Scientific, A-21436) (Diluted 1 to 1000)

Anti-Mouse DyLight 488 made in Donkey (Thermo Fisher Scientific, SA5-10166) (Diluted 1 to 400)

Anti-Rabbit DyLight 650 made in Donkey (Thermo Fisher Scientific, SA5-10041) (Diluted 1 to 400)

Solutions:

Washing Solution: PBS

Permeabilization Solution: PBS with 0.3% Triton X-100 (PBST) (Sigma Aldrich, 9002-93-1)

Blocking Solution: PBST with 1% BSA (PBSTb) (CHEM-IMPEX INT’L, 00535)

Sperm: Frozen human and cattle sperm were stored in liquid nitrogen and obtained from UToledo andrology clinic and Select Sires Inc. IRB number 202366-UT and 300220-UT, and IBC number108074-UT

Cattle sperm was removed from the straws and human sperm was removed from cryovials. Both types of samples were put onto slides in 5 steps: 1. Straw or cryovials were placed in 37°C water; 2. Sperm was washed using a 40/80 density gradient (Nidacon, Cat # PS40-100 and PS80-100); 3. The pellet was collected and washed in sperm washing media (Nidacon, Cat # PSW-100), then resuspended in M199 (Sigma-Aldrich, Cat # M7528-500ML); 5. Sperm was added to the slide and then dropped into liquid nitrogen where it was stored until use.

Fixation: Methanol (-20°C) (Fisher Chemical, A412P-4)

A humidity chamber is a plastic container that is layered with wet paper towels covered with a lid

Material:

Confocal Microscope: Leica SP8, Sperm images were taken at a magnification of 640× and zoom of 6×, with 512 × 512-pixel density, 3x zoom and 1024x1024-pixel density or 0.75x zoom and 4096x4096-pixel density

Using Photoshop, immunofluorescence sperm panels were cropped to:

- Low magnification: 525 x 1050 pixels and adjusted to 1-inch x 2 inches

- Medium magnification: 150 x 300 pixels and adjusted to 1-inch x 2 inches

- High magnification: 75 x 75 pixels and adjusted to 1-inch x 1 inch.

Adobe Illustrator was used to create sperm images.

Parafilm Wax (VWR, 52858-032)

Cover Slip (VWR, 48366-205)

Slides (Home Science Tools, MS-SLFRO72)

Nail Polish (EMS Diasum, 72180)

Mounting Media made of Fluoroshield with DAPI (Sigma-Aldrich, F6057-20ML)

Hoechst (Thermo Fisher Scientific, H1399) (Diluted 1 to 2000)

LAS X Software Leica using photon counting and BrightR.

Statistical methods: Experiments were completed 2-4 times; we calculated averages and standard deviations using Microsoft Excel.

Acknowledgments

Acknowledgments

We would like to thank Rebecca Wynn for collecting sperm samples.

Funding

This project was supported by Agriculture and Food Research Initiative Competitive Grant no. OHOW-2020-02790 from the USDA National Institute of Food and Agriculture. JS and JT were supported by the MSRP at the University of Toledo College of Medicine and Life Sciences

References

  1. Amargant F, Pujol A, Ferrer-Vaquer A, Durban M, Martínez M, Vassena R, Vernos I. The human sperm basal body is a complex centrosome important for embryo preimplantation development. Mol Hum Reprod. 2021 Nov 2;27(11) doi: 10.1093/molehr/gaab062. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Arnold J, Schattschneider J, Blechner C, Krisp C, Schlüter H, Schweizer M, Nalaskowski M, Oliveira-Ferrer L, Windhorst S. Tubulin Tyrosine Ligase Like 4 (TTLL4) overexpression in breast cancer cells is associated with brain metastasis and alters exosome biogenesis. J Exp Clin Cancer Res. 2020 Sep 30;39(1):205–205. doi: 10.1186/s13046-020-01712-w. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bobinnec Y, Moudjou M, Fouquet JP, Desbruyères E, Eddé B, Bornens M. Glutamylation of centriole and cytoplasmic tubulin in proliferating non-neuronal cells. Cell Motil Cytoskeleton. 1998;39(3):223–232. doi: 10.1002/(SICI)1097-0169(1998)39:3<223::AID-CM5>3.0.CO;2-5. [DOI] [PubMed] [Google Scholar]
  4. Bowker Z, Goldstein S, Breitbart H. Protein acetylation protects sperm from spontaneous acrosome reaction. Theriogenology. 2022 Aug 12;191:231–238. doi: 10.1016/j.theriogenology.2022.08.005. [DOI] [PubMed] [Google Scholar]
  5. Cavazza T, Takeda Y, Politi AZ, Aushev M, Aldag P, Baker C, Choudhary M, Bucevičius J, Lukinavičius G, Elder K, Blayney M, Lucas-Hahn A, Niemann H, Herbert M, Schuh M. Parental genome unification is highly error-prone in mammalian embryos. Cell. 2021 May 7;184(11):2860–2877.e22. doi: 10.1016/j.cell.2021.04.013. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chang CW, Hsu WB, Tsai JJ, Tang CJ, Tang TK. CEP295 interacts with microtubules and is required for centriole elongation. J Cell Sci. 2016 May 16;129(13):2501–2513. doi: 10.1242/jcs.186338. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chawan V, Yevate S, Gajbhiye R, Kulkarni V, Parte P. Acetylation/deacetylation and microtubule associated proteins influence flagellar axonemal stability and sperm motility. Biosci Rep. 2020 Dec 23;40(12) doi: 10.1042/BSR20202442. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Chemes HE, Alvarez Sedo C. Tales of the tail and sperm head aches: changing concepts on the prognostic significance of sperm pathologies affecting the head, neck and tail. Asian J Androl. 2011 Dec 26;14(1):14–23. doi: 10.1038/aja.2011.168. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Eddé B, Rossier J, Le Caer JP, Desbruyères E, Gros F, Denoulet P. Posttranslational glutamylation of alpha-tubulin. Science. 1990 Jan 5;247(4938):83–85. doi: 10.1126/science.1967194. [DOI] [PubMed] [Google Scholar]
  10. Eshun-Wilson L, Zhang R, Portran D, Nachury MV, Toso DB, Löhr T, Vendruscolo M, Bonomi M, Fraser JS, Nogales E. Effects of α-tubulin acetylation on microtubule structure and stability. Proc Natl Acad Sci U S A. 2019 May 9;116(21):10366–10371. doi: 10.1073/pnas.1900441116. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Fishman EL, Jo K, Nguyen QPH, Kong D, Royfman R, Cekic AR, Khanal S, Miller AL, Simerly C, Schatten G, Loncarek J, Mennella V, Avidor-Reiss T. A novel atypical sperm centriole is functional during human fertilization. Nat Commun. 2018 Jun 7;9(1):2210–2210. doi: 10.1038/s41467-018-04678-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fouquet JP, Edde B, Kann ML, Wolff A, Desbruyeres E, Denoulet P. Differential distribution of glutamylated tubulin during spermatogenesis in mammalian testis. Cell Motil Cytoskeleton. 1994;27(1):49–58. doi: 10.1002/cm.970270106. [DOI] [PubMed] [Google Scholar]
  13. Gadadhar S, Alvarez Viar G, Hansen JN, Gong A, Kostarev A, Ialy-Radio C, Leboucher S, Whitfield M, Ziyyat A, Touré A, Alvarez L, Pigino G, Janke C. Tubulin glycylation controls axonemal dynein activity, flagellar beat, and male fertility. Science. 2021 Jan 8;371(6525) doi: 10.1126/science.abd4914. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Gadadhar S, Dadi H, Bodakuntla S, Schnitzler A, Bièche I, Rusconi F, Janke C. Tubulin glycylation controls primary cilia length. J Cell Biol. 2017 Jul 7;216(9):2701–2713. doi: 10.1083/jcb.201612050. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hamel V, Steib E, Hamelin R, Armand F, Borgers S, Flückiger I, Busso C, Olieric N, Sorzano COS, Steinmetz MO, Guichard P, Gönczy P. Identification of Chlamydomonas Central Core Centriolar Proteins Reveals a Role for Human WDR90 in Ciliogenesis. Curr Biol. 2017 Aug 3;27(16):2486–2498.e6. doi: 10.1016/j.cub.2017.07.011. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Jaiswal A, Baliu-Souza T, Turner K, Nadiminty N, Rambhatla A, Agarwal A, Krawetz SA, Dupree JM, Saltzman B, Schon SB, Avidor-Reiss T. Sperm centriole assessment identifies male factor infertility in couples with unexplained infertility - a pilot study. Eur J Cell Biol. 2022 May 27;101(3):151243–151243. doi: 10.1016/j.ejcb.2022.151243. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Janke C, Magiera MM. The tubulin code and its role in controlling microtubule properties and functions. Nat Rev Mol Cell Biol. 2020 Feb 27;21(6):307–326. doi: 10.1038/s41580-020-0214-3. [DOI] [PubMed] [Google Scholar]
  18. Kalebic N, Sorrentino S, Perlas E, Bolasco G, Martinez C, Heppenstall PA. αTAT1 is the major α-tubulin acetyltransferase in mice. Nat Commun. 2013;4:1962–1962. doi: 10.1038/ncomms2962. [DOI] [PubMed] [Google Scholar]
  19. Kann ML, Prigent Y, Levilliers N, Bré MH, Fouquet JP. Expression of glycylated tubulin during the differentiation of spermatozoa in mammals. Cell Motil Cytoskeleton. 1998;41(4):341–352. doi: 10.1002/(SICI)1097-0169(1998)41:4<341::AID-CM6>3.0.CO;2-8. [DOI] [PubMed] [Google Scholar]
  20. Khanal S, Leung MR, Royfman A, Fishman EL, Saltzman B, Bloomfield-Gadêlha H, Zeev-Ben-Mordehai T, Avidor-Reiss T. A dynamic basal complex modulates mammalian sperm movement. Nat Commun. 2021 Jun 21;12(1):3808–3808. doi: 10.1038/s41467-021-24011-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. LeDizet M, Piperno G. Identification of an acetylation site of Chlamydomonas alpha-tubulin. Proc Natl Acad Sci U S A. 1987 Aug 1;84(16):5720–5724. doi: 10.1073/pnas.84.16.5720. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Liu Z, Nguyen QPH, Nanjundappa R, Delgehyr N, Megherbi A, Doherty R, Thompson J, Jackson C, Albulescu A, Heng YM, Lucas JS, Dell SD, Meunier A, Czymmek K, Mahjoub MR, Mennella V. Super-Resolution Microscopy and FIB-SEM Imaging Reveal Parental Centriole-Derived, Hybrid Cilium in Mammalian Multiciliated Cells. Dev Cell. 2020 Oct 9;55(2):224–236.e6. doi: 10.1016/j.devcel.2020.09.016. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Lobert VH, Skardal ML, Malerød L, Simensen JE, Algra HA, Andersen AN, Fleischer T, Enserink HA, Liestøl K, Heath JK, Rusten TE, Stenmark HA. PHLPP1 regulates CFTR activity and lumen expansion through AMPK. Development. 2022 Aug 23;149(20) doi: 10.1242/dev.200955. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Magiera MM, Janke C. Post-translational modifications of tubulin. Curr Biol. 2014 May 5;24(9):R351–R354. doi: 10.1016/j.cub.2014.03.032. [DOI] [PubMed] [Google Scholar]
  25. Mahecic D, Gambarotto D, Douglass KM, Fortun D, Banterle N, Ibrahim KA, Le Guennec M, Gönczy P, Hamel V, Guichard P, Manley S. Homogeneous multifocal excitation for high-throughput super-resolution imaging. Nat Methods. 2020 Jun 22;17(7):726–733. doi: 10.1038/s41592-020-0859-z. [DOI] [PubMed] [Google Scholar]
  26. Nogales E, Whittaker M, Milligan RA, Downing KH. High-resolution model of the microtubule. Cell. 1999 Jan 8;96(1):79–88. doi: 10.1016/s0092-8674(00)80961-7. [DOI] [PubMed] [Google Scholar]
  27. Pandruvada S, Royfman R, Shah TA, Sindhwani P, Dupree JM, Schon S, Avidor-Reiss T. Lack of trusted diagnostic tools for undetermined male infertility. J Assist Reprod Genet. 2021 Jan 2;38(2):265–276. doi: 10.1007/s10815-020-02037-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Piperno G, LeDizet M, Chang XJ. Microtubules containing acetylated alpha-tubulin in mammalian cells in culture. J Cell Biol. 1987 Feb 1;104(2):289–302. doi: 10.1083/jcb.104.2.289. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Piroli GG, Manuel AM, Walla MD, Jepson MJ, Brock JW, Rajesh MP, Tanis RM, Cotham WE, Frizzell N. Identification of protein succination as a novel modification of tubulin. Biochem J. 2014 Sep 1;462(2):231–245. doi: 10.1042/BJ20131581. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Portran D, Schaedel L, Xu Z, Théry M, Nachury MV. Tubulin acetylation protects long-lived microtubules against mechanical ageing. Nat Cell Biol. 2017 Feb 27;19(4):391–398. doi: 10.1038/ncb3481. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Redeker V, Levilliers N, Schmitter JM, Le Caer JP, Rossier J, Adoutte A, Bré MH. Polyglycylation of tubulin: a posttranslational modification in axonemal microtubules. Science. 1994 Dec 9;266(5191):1688–1691. doi: 10.1126/science.7992051. [DOI] [PubMed] [Google Scholar]
  32. Reerink KE. Quality assurance in health care of developing countries. Qual Assur Health Care. 1989;1(4):195–196. [PubMed] [Google Scholar]
  33. Soppina V, Herbstman JF, Skiniotis G, Verhey KJ. Luminal localization of α-tubulin K40 acetylation by cryo-EM analysis of fab-labeled microtubules. PLoS One. 2012 Oct 26;7(10):e48204–e48204. doi: 10.1371/journal.pone.0048204. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Sullenberger C, Vasquez-Limeta A, Kong D, Loncarek J. With Age Comes Maturity: Biochemical and Structural Transformation of a Human Centriole in the Making. Cells. 2020 Jun 9;9(6) doi: 10.3390/cells9061429. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Terada Y, Nakamura S, Simerly C, Hewitson L, Murakami T, Yaegashi N, Okamura K, Schatten G. Centrosomal function assessment in human sperm using heterologous ICSI with rabbit eggs: a new male factor infertility assay. Mol Reprod Dev. 2004 Mar 1;67(3):360–365. doi: 10.1002/mrd.20024. [DOI] [PubMed] [Google Scholar]
  36. Thazhath R, Jerka-Dziadosz M, Duan J, Wloga D, Gorovsky MA, Frankel J, Gaertig J. Cell context-specific effects of the beta-tubulin glycylation domain on assembly and size of microtubular organelles. Mol Biol Cell. 2004 Jul 14;15(9):4136–4147. doi: 10.1091/mbc.e04-03-0247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Thoma ME, McLain AC, Louis JF, King RB, Trumble AC, Sundaram R, Buck Louis GM. Prevalence of infertility in the United States as estimated by the current duration approach and a traditional constructed approach. Fertil Steril. 2013 Jan 3;99(5):1324–1331.e1. doi: 10.1016/j.fertnstert.2012.11.037. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Tremoleda JL, Van Haeften T, Stout TA, Colenbrander B, Bevers MM. Cytoskeleton and chromatin reorganization in horse oocytes following intracytoplasmic sperm injection: patterns associated with normal and defective fertilization. Biol Reprod. 2003 Mar 19;69(1):186–194. doi: 10.1095/biolreprod.102.012823. [DOI] [PubMed] [Google Scholar]
  39. Turner KA, Fishman EL, Asadullah M, Ott B, Dusza P, Shah TA, Sindhwani P, Nadiminty N, Molinari E, Patrizio P, Saltzman BS, Avidor-Reiss T. Fluorescence-Based Ratiometric Analysis of Sperm Centrioles (FRAC) Finds Patient Age and Sperm Morphology Are Associated With Centriole Quality. Front Cell Dev Biol. 2021 Apr 22;9:658891–658891. doi: 10.3389/fcell.2021.658891. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. van Dijk J, Rogowski K, Miro J, Lacroix B, Eddé B, Janke C. A targeted multienzyme mechanism for selective microtubule polyglutamylation. Mol Cell. 2007 May 11;26(3):437–448. doi: 10.1016/j.molcel.2007.04.012. [DOI] [PubMed] [Google Scholar]
  41. Vogel P, Hansen G, Fontenot G, Read R. Tubulin tyrosine ligase-like 1 deficiency results in chronic rhinosinusitis and abnormal development of spermatid flagella in mice. Vet Pathol. 2010 May 4;47(4):703–712. doi: 10.1177/0300985810363485. [DOI] [PubMed] [Google Scholar]
  42. Wloga D, Webster DM, Rogowski K, Bré MH, Levilliers N, Jerka-Dziadosz M, Janke C, Dougan ST, Gaertig J. TTLL3 Is a tubulin glycine ligase that regulates the assembly of cilia. Dev Cell. 2009 Jun 1;16(6):867–876. doi: 10.1016/j.devcel.2009.04.008. [DOI] [PubMed] [Google Scholar]
  43. Xu Z, Schaedel L, Portran D, Aguilar A, Gaillard J, Marinkovich MP, Théry M, Nachury MV. Microtubules acquire resistance from mechanical breakage through intralumenal acetylation. Science. 2017 Apr 21;356(6335):328–332. doi: 10.1126/science.aai8764. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from microPublication Biology are provided here courtesy of California Institute of Technology

RESOURCES