The Cerebellum

https://doi.org/10.1007/s12311-020-01167-x

SHORT REPORTS

Factors Associated with Intergenerational Instability of ATXN3 CAG

Repeat and Genetic Anticipation in Chinese Patients
with Spinocerebellar Ataxia Type 3
Yi-Chu Du 1 & Yin Ma 1 & Ya-Ru Shao 1 & Shi-Rui Gan 2 & Yi Dong 1,3 & Zhi-Ying Wu 1

# Springer Science+Business Media, LLC, part of Springer Nature 2020

Abstract
Spinocerebellar ataxia type 3 (SCA3) is caused by unstable expanded CAG repeats (expCAGs) in ATXN3. Factors associated
with intergenerational instability (delta-expCAG) and genetic anticipation in SCA3 have never been reported in Chinese main-
land. Here, we demonstrated that unstable transmissions occurred more often in sons than in daughters (91% vs 72%, Fisher’s
exact test, p = 0.012). The extended delta-expCAG of father-son transmissions was greater than that of mother-son transmissions
(3.8 ± 2.3 repeats vs 1.6 ± 1.0 repeats, Mann-Whitney U, p = 0.001). Genetic anticipation was frequently observed between
generations but not affected by the delta-expCAG.

Keywords Spinocerebellar ataxia type 3 . Intergenerational transmission . Genetic anticipation . Chinese

Introduction genetic anticipation [5]. Revealing the factors associated with

Spinocerebellar ataxia type 3 (SCA3) is the most common
subtype of spinocerebellar ataxias (SCAs) in the Chinese pop-
ulation. It is an autosomal dominant neurodegenerative dis-
ease caused by an unstable expansion of CAG repeats in
ATXN3 [1]. The age at onset (AAO) of SCA3 is mostly mod-
ified by the size of the expanded CAG repeats (expCAGs).
During intergenerational transmission, the expCAG instability
and genetic anticipation are the features in polyglutamine
(polyQ) diseases such as Huntington’s disease (HD) and
SCAs [2–4]. Moreover, this transmitted instability (i.e., del-
ta-expCAG) is thought to explain the phenotype, such as
Yi-Chu Du and Yin Ma contributed equally to this work.
* Zhi-Ying Wu
zhiyingwu@zju.edu.cn
CAG instability and genetic anticipation would be helpful in
clinical practice in SCA3 and helpful to unveil underlying
mechanism in polyQ diseases.
Indeed, factors affecting the transmitted instability in
polyQ disease remain inconsistent. More factors associated
with the transmitted instability are still under research [6, 7,
8]. As genotype in SCA3 varied in different populations, in-
vestigations of affecting factors in different populations are of
great vital. The associated factors of transmitted instability
have been reported in Japanese, Brazilian, and Portuguese
[5, 7, 9], but few have been documented in Chinese except
Chinese people in Taiwan [ 10, 11]. Here, we aim to investi-
gate the factors associated with intergenerational instability of
CAG repeats and genetic anticipation in mainland Chinese
SCA3 patients.

1
Department of Neurology and Research Center of Neurology in Materials and Methods
Second Affiliated Hospital, and Key Laboratory of Medical
Neurobiology of Zhejiang Province, Zhejiang University School of
Medicine, Hangzhou 310009, China Participants for diagnosis and/or genetic counseling were re-
2
Department of Neurology and Institute of Neurology, First Affiliated
cruited between July 1, 2009, and September 10, 2019, from
Hospital, Fujian Medical University, Fuzhou 350005, China Departments of Neurology in three leading academic hospi-
3
Department of Neurology and Institute of Neurology, Huashan
tals, including the Second Affiliated Hospital of Zhejiang
Hospital, Shanghai Medical College, Fudan University, University (Hangzhou), the First Affiliated Hospital of
Shanghai 200041, China Fujian Medical University (Fuzhou), and Huashan Hospital

of Fudan University (Shanghai). These participants partly o-
verlapped with the subjects in previous research for different
purposes [12]. Peripheral blood samples were collected for
sequencing CAG repeat as previously reported [13]. The val-
ue of delta-expCAG is the expCAG change which occurs
between the offspring and the parents during the germline
transmission. Stable transmission is defined by a delta-
expCAG equal to 0; otherwise, it is extended or contracted
transmission. Genetic anticipation is the value of delta-AAO,
and positive value means the offspring has an earlier AAO
than the parent. Age of conception is the age of the transmit-
ting parent at conception. Statistical analysis was performed
using SPSS 20.0 and Microsoft Excel 2016. The paired t test,
Mann-Whitney U test, nonparametric correlation, or partial
correlation was used when appropriate. It was considered sig-
nificant difference when a p value < 0.05. Since offspring’s
AAO significantly correlated to genetic anticipation and off-
spring’s expCAG, we excluded the effect of offspring’s AAO
when evaluating the relationship between genetic anticipation
and offspring’s expCAG.
Results
During the last 10 years, the delta-expCAG was obtained from
91 SCA3 families involving 112 parent-child pairs (54 daugh-
ters and 58 sons), and genetic anticipation was obtained from
133 families including 135 SCA3 patients (80 females and 55
males). In 112 parent-child transmissions (46 paternal and 66
maternal), the mean size of the expCAG among offspring was
significantly larger than that among their parents (paired t test,
p < 0.001), with a delta-expCAG of 1.0 ± 2.6 repeats (range −
11 to 9). The distribution of 112 delta-expCAGs is depicted in
Fig. 1a. Ninety-two of 112 (82.1%) pairs presented with the
offspring having unstable transmissions (66 had expansions
and 26 had contractions), while 20 pairs remained unchanged
(Table 1). As shown in Table 1, the mean age of conception
was 25.6 ± 3.8 years (range 19–47 years) in 112 parent-child
transmissions, while it was 27.2 ± 4.2 years and 24.6 ±
3.1 years in 46 paternal transmissions and 66 maternal trans-
missions, respectively. Genetic anticipation from 135 patients
Fig. 1 Distribution of intergenerational instability of delta-expCAG and
genetic anticipation during germline transmissions. a Distribution of
delta-expCAG at the pathogenic allele in maternal and paternal transmis-
sions from 112 SCA3 parent-child pairs. b Distribution of maternal and
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(75 paternal transmissions and 60 maternal transmissions) was
observed 9.2 ± 9.4 years (range − 31 to 35 years) earlier
among offspring than among their parents. The distribution
of genetic anticipation is depicted in Fig. 1b. Among 135
SCA3 patients, 113 (83.7%) had an earlier AAO than their
parents; however, 17 (12.6%) had a later AAO and 5 (3.7%)
had an unchanged AAO.
We firstly investigated the factors associated with intergen-
erational CAG repeat instability. We focused on whether the
portion of unstable transmissions, extended/contracted trans-
missions, and the delta-expCAG were associated with any of
those affecting factors, including parental and offspring’s sex,
parental expCAG, and age at conception. Unexpectedly, we
found that unstable transmissions happened more often in
sons than in daughters (91% vs 72%, Fisher’s exact test, p =
0.012). No difference of unstable transmission was observed
either between paternal-son and paternal-daughter transmis-
sions (Fisher’s exact test, p = 0.07) or between maternal-son
and maternal-daughter transmissions (Fisher’s exact test, p =
0.13). In addition, neither portion of unstable transmissions
nor contracted/extended transmissions were related to parental
sex, parental expCAG, or age at conception.
As shown in Table 1, the mean delta-expCAG was larg-
er in 46 paternal transmissions (1.4 ± 3.0 repeats) than in
66 maternal transmissions (0.7 ± 2.1 repeats), although not
statistically significant (Mann-Whitney U, p = 0.64). The
discrepancy of delta-expCAG between paternal transmis-
sion (3.5 ± 2.4 repeats) and maternal transmission (1.9 ±
1.2 repeats) became more evident (Mann-Whitney U, p =
0.004) when only extended transmission was considered.
Moreover, we observed that sons with paternal transmis-
sion had the largest delta-expCAG (2.2 ± 3.2 repeats) and
the largest extended delta-expCAG (3.8 ± 2.3 repeats),
whereas daughters with maternal transmission had the
greatest contracted delta-expCAG (− 3.8 ± 4.9 repeats).
However, only the extended delta-expCAG between
father-son transmissions (3.8 ± 2.3 repeats) and mother-
son transmissions (1.6 ± 1.0 repeats) was detected signifi-
cantly different (Mann-Whitney U, p = 0.001).
Additionally, the delta-expCAG was explained by neither
age at conception nor parental expCAG.
paternal anticipation in 135 SCA3 patients. c Relationship between an-
ticipation and delta-expCAG in 9 SCA3 maternal and 9 paternal
transmissions
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Table 1 Intergenerational transmissions in Chinese SCA3 families

Total Paternal Maternal

Total Daughters Sons Total Daughters Sons

Transmissions, n (%) 112 46 (41.1) 20 (17.9) 26 (23.1) 66 (58.9) 34 (30.4) 32 (28.6)

Unstable 92 (82.1) 40 (86.9) 15 (75.0) 25 (96.2) 52 (78.8) 24 (70.6) 28 (87.5)
transmissions, n (%)
Contractions, n (%) 26 (23.2) 14 (30.4) 7 (35.0) 7 (26.9) 12 (18.2) 4 (11.8) 8 (25.0)
Expansions, n (%) 66 (58.9) 26 (56.5) 8 (40.0) 18 (69.2) 40 (60.6) 20 (58.8) 20 (62.5)
Parental expCAG 73.4 ± 3.2 73.6 ± 3.1 74.2 ± 3.4 73.2 ± 2.8 73.3 ± 3.3 73.4 ± 2.7 73.2 ± 4.0
Mean ± SD (range) (58 to 82) (66 to 82) (68 to 82) (66 to 77) (58 to 80) (67 to 77) (58 to 80)
Delta-expCAG 1.0 ± 2.6 1.4 ± 3.0 0.5 ± 2.6 2.2 ± 3.2 0.7 ± 2.1 0.8 ± 2.6 0.7 ± 1.5
Mean ± SD (range) (− 11 to 9) (− 4 to 9) (− 4 to 7) (− 3 to 9) (− 11 to 5) (− 11 to 5) (− 2 to 4)
Contracted − 2.0 ± 2.0 − 1.8 ± 1.0 − 1.9 ± 1.1 − 1.7 ± 1.0 − 2.2 ± 2.8 − 3.8 ± 4.9 − 1.4 ± 0.5
delta-expCAG (− 11 to − 1) (− 4 to − 1) (− 4 to − 1) (− 3 to − 1) (− 11 to − 1) (− 11 to −1) (− 2 to − 1)
Mean ± SD (range)
Extended 2.5 ± 1.9 3.5 ± 2.4 a 2.8 ± 2.4 3.8 ± 2.3b 1.9 ± 1.2 a 2.1 ± 1.2 1.6 ± 1.0 b
delta-expCAG (1 to 9) (1 to 9) (1 to 7) (1 to 9) (1 to 5) (1 to 5) (1 to 4)
Mean ± SD (range)
Age at conception 25.6 ± 3.8 27.2 ± 4.2 24.6 ± 3.1
Mean ± SD (range) (19 to 47) (22 to 47) (19 to 36)
Anticipation, n (%) 135 75 (55.6) 50 (37.0) 25 (18.6) 60 (44.4) 30 (22.2) 30 (22.2)
Mean ± SD (range) 9.2 ± 9.4 (− 31 10.2 ± 10.4 (− 31 9.9 ± 8.8 (− 10 10.8 ± 13.3 (− 31 7.9 ± 7.8 (− 8 7.8 ± 7.2 (− 2 8.1 ± 8.5 (− 8
to 35) to 35) to 24) to 35) to 26) to 24) to 26)
a
Compared to maternal transmissions, extended delta-expCAG was larger in paternal transmissions, Mann-Whitney U, p = 0.004
b
Compared to maternal sons, extended delta-expCAG was larger in paternal sons, Mann-Whitney U, p = 0.001
n, number; expCAG, expanded CAG repeat

Then we investigated the factors associated with genetic
anticipation which were not previously reported in China.
The factors included age at conception, parental and off-
spring’s sex, parental and offspring’s expCAG, and delta-
expCAG. Genetic anticipation of 135 patients was 9.2 ±
9.4 years and showed no significant difference either between
paternal and maternal transmissions (Table 1, Mann-Whitney
U, p = 0.053) or between male (9.1 ± 8.3 years) and female
patients (9.3 ± 10.9 years, t test, p = 0.9). Eighteen out of
135 patients had the age at conception (25.3 ± 2.1, 22–
30 years), parental expCAG (74.2 ± 3.0, 67–82 repeats), and
delta-expCAG (3.2 ± 3.0, − 2 to 9 repeats). The delta-expCAG
showed no association with anticipation (16.8 ± 7.7, 0–35)
among the 18 patients regardless of whether parental sex
was considered (Fig. 1c, nonparametric correlation, each
p > 0.4). Additionally, neither of age at conception and paren-
tal expCAG was associated with genetic anticipation (non-
parametric correlation, each p > 0.05), nor was offspring’s
expCAG (partial correlation, p = 0.56).
Discussion
Transmitted instability and genetic anticipation were rarely
reported in the Chinese SCA3 pedigrees. Only 14 and 16
parent-child pairs of SCA3 were, respectively, reported in
Chinese people in Taiwan [10, 11]. In Soong’s report [10],
64.3% of 14 pairs were unstable transmissions with a delta-
expCAG of 1.5 repeats, while in Hsieh’s report [11], the delta-
expCAG was 1.8 repeats. Here in the current study, we dem-
onstrated a delta-expCAG of 1.0 repeat and a higher percent-
age of unstable transmissions (82.1%) than Soong’s report
(64.3%) [10] and Portuguese report (71%) [7] but similar to
a Brazilian research (81.8%) [5]. Whereas in Japan, Brazil,
and Portugal, mean delta-expCAG was 1.9 repeats (n = 30),
1.7 repeats (n = 115), and 0.86 repeats (n = 100), respectively
[5, 7, 9].
Notably, we found that the portion of unstable transmis-
sions was related to the offspring’s sex instead of the parental
sex, which is scarcely reported in SCA3. It is still commonly
believed that paternal sex showed great influence on the CAG
instability in SCA3 [2]. However in HD, the direction and the
value of the delta-expCAG were reported dependent upon the
offspring’s sex during maternal transmissions [14]. Although
we did not find offspring’s sex mattered during maternal trans-
missions, we found offspring’s sex had an impact on both
delta-expCAG and portion of unstable transmissions.
Additionally, direct correlation between parental sex and
delta-expCAG is less prominent in the current study, where
the greater instability of paternal transmission in SCA3 could
only be observed in the extended delta-expCAG group rather
than the total delta-expCAG group. Even though as reported,

paternal transmission was more unstable not only in SCA3 [2]
but in many other polyQ diseases such as SCA2 and HD [3,
4]. Along with the result that larger extended delta-expCAG
was observed in paternal-son transmission than maternal-son
transmission, it might implicate that offspring’s sex is worth
more notices, where the importance of offspring’s sex was
underestimated in SCA3.
Currently, we did not find the correlation between the
delta-expCAG and parental expCAGs in the current study,
confirming the previous findings in SCA3 [1, 5]. The greater
instability was reported to increase with parental age of con-
ception in both SCA2 and SCA3 [4, 5, 7], which might indi-
cate aging is functioning in meiotic instability since aged par-
ents delivered more unstable disease alleles to their offspring.
However, it is still arguable as the contradictory findings ob-
tained from the current study and previous HD studies [3,14],
where no correlation between CAG instability and age of con-
ception is observed. Research has been exhibiting more pos-
sibilities to explain CAG instability in SCA3. Variants at
DNA replication, repair and recombination genes, and DNA
methylation levels of the promotor at ATXN3 gene were found
to be associated with CAG instability in SCA3 [7, 15].
In addition, we detected severe anticipation with small
delta-expCAG in a subgroup of 18 parent-child pairs here,
and no relationship between genetic anticipation and delta-
expCAG was observed. Contrarily, it was reported that the
genetic anticipation was correlated with delta-expCAG (r =
0.38, 74 pairs) in Brazil; however, the correlation was much
weaker than that between AAO and the expCAG (r = − 0.80)
[5]. Therefore, the current study and Brazilian study both in-
dicate that there probably are more unclear impactors affecting
genetic anticipation. However, current sample size is still lim-
ited, which might lead to the observation bias. Therefore, the
investigation involving more patients should be well merited.
Conclusion
Our study demonstrates the first investigation of transmission
mode and the associated factors in the largest mainland
Chinese SCA3 families. Transmitted offspring’s sex partici-
pated in the delta-expCAG and portion of unstable transmis-
sions. Genetic anticipation was not related to any factor. These
findings will deepen the understanding of intergenerational
transmissions in SCA3 and be useful for future research on
polyQ diseases.
Acknowledgments We would like to thank all participants for their will-
ingness to join this study.
Authors’ Contribution Yi-Chu Du: data collection, analysis and interpre-
tation, and draft of the manuscript.
Yin Ma: administrative and technical support and data analysis and
interpretation.
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Ya-Ru Shao: data collection.
Shi-Rui Gan: data collection.
Yi Dong: data collection and analysis and interpretation.
Zhi-Ying Wu: study concept and design, data acquiring, analysis and
interpretation, and critical revision of the manuscript.
Funding Information This work was supported by the research founda-
tion for distinguished scholar of Zhejiang University to Zhi-Ying Wu
(188020-193810101/089, Hangzhou).
Data Availability The data that support the findings of this study are
available from the corresponding author upon reasonable request.
Compliance with Ethical Standards
Conflict of Interest The authors declare that they have no conflict of
interest.
Ethics Approval This study was performed in line with the principles of
the Declaration of Helsinki. And the study was approved by the ethics
committees of the Second Affiliated Hospital of Zhejiang University
(Hangzhou), the First Affiliated Hospital of Fujian Medical University
(Fuzhou), and Huashan Hospital of Fudan University (Shanghai).
Consent to Participate Written informed consent was obtained from
each participant.
Consent to Publish Not applicable.
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