Screening and functional analysis of proteins interacting with chicken adenylosuccinate lyase

doi:10.3969/j.issn.1004-1524.20230237

Acta Agriculturae Zhejiangensis ›› 2024, Vol. 36 ›› Issue (3): 515-526.DOI: 10.3969/j.issn.1004-1524.20230237

• Animal Science • Previous Articles Next Articles

Screening and functional analysis of proteins interacting with chicken adenylosuccinate lyase

YU Huan(), LI Hui(), CHEN Youbo, SHI Yushi, ZHAO Depeng, LONG Xia, TAN Qisong

Key Laboratory of Genetic Breeding and Reproduction of Plateau and Mountain Animals, Ministry of Education, College of Animal Science, Guizhou University, Guiyang 550025, China

Received:2023-03-01 Online:2024-03-25 Published:2024-04-09

Abstract

Abstract:

By exploring the regulatory relationship between chicken adenylosuccinate lyase gene ADSL and its interaction genes, to provide reference for further study on the reason of high expression of ADSL in muscle and the mechanism of ADSL affecting muscle flavor. In this study, the recombinant eukaryotic expression vector pEGFP-C1-ADSL was constructed. The pEGFP-C1-ADSL and pEGFP-C1 plasmids were transfected into chicken myoblasts respectively, and the total protein of the successfully expressed cells was extracted. The cellular proteins interacting with chicken ADSL protein were identified by co-immunoprecipitation (Co-IP ) combined with mass spectrometry, and the results were analyzed by GO functional annotation, KEGG pathway and protein interaction network. RPL4, PDLIM5, ACTG1 and SRSF10 were screened, and the expression of RPL4, PDLIM5, ACTG1 and SRSF10 genes in myoblasts were detected under overexpression and silencing of ADSL gene. The results showed that 94 proteins interacted with ADSL. Bioinformatics analysis showed that these proteins were localized to cellular anatomical entity, intracellular and protein-containing complexs, and they were mainly involved in biological processes such as cellular processes, biological regulation, and stimulus responses. The results of indirect immunofluorescence showed that the recombinant protein pEGFP-C1-ADSL was mainly localized in the nucleus and cytoplasm. When ADSL was overexpressed, the expression of RPL4 and PDLIM5 genes in myoblasts was down-regulated, and the expression of ACTG1 and SRSF10 genes was up-regulated. When ADSL was silenced, the expression of RPL4 and ACTG1 genes in myoblasts was up-regulated. The results enriched the ADSL protein that regulates flavor, and provided a reference for further understanding the function of ADSL and the high expression of this gene in muscle.

Key words: adenylosuccinate lyase, Chishui black bone chicken, immunoprecipitation, interacting protein

CLC Number:

S834.83

YU Huan, LI Hui, CHEN Youbo, SHI Yushi, ZHAO Depeng, LONG Xia, TAN Qisong. Screening and functional analysis of proteins interacting with chicken adenylosuccinate lyase[J]. Acta Agriculturae Zhejiangensis, 2024, 36(3): 515-526.

Figures/Tables 10

References 28

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UniProt登录号	蛋白质名称	基因名	长度	分子量	覆盖度
UniProt accession	Protein name	Gene name	Length/aa	Mass/ku	Coverage/%
P97461	核糖体蛋白S5 Ribosomal protein S5	Rps5	204	22.9	4.41
Q9CZX8	核糖体蛋白S19 Ribosomal protein S19	Rps19	145	16.1	8.97
P60867	核糖体蛋白S20 Ribosomal protein S20	Rps20	119	13.4	10.08
P62858	核糖体蛋白S28 Ribosomal protein S28	Rps28	69	7.8	17.39
P62274	核糖体蛋白S29 Ribosomal protein S29	Rps29	56	6.7	14.29
Q9D8E6	核糖体蛋白L4 Ribosomal protein L4	Rpl4	419	47.2	12.89
P47962	核糖体蛋白L5 Ribosomal protein L5	Rpl5	297	34.4	12.79
O09167	核糖体蛋白L21 Ribosomal protein L21	Rpl21	160	18.6	6.88
P61255	核糖体蛋白L26 Ribosomal protein L26	Rpl26	145	17.3	9.66
P62900	核糖体蛋白L31 Ribosomal protein L31	Rpl31	125	14.5	18.40
P62717	核糖体蛋白L18A Ribosomal protein L18A	Rpl18a	176	20.7	15.34
P99027	核糖体蛋白,大P2 Ribosomal protein, large P2	Rplp2	115	11.7	25.22
Q7TPR4	肌动蛋白1 Actinin 1	Actn1	892	103.1	4.04
P68134	肌动蛋白α1 Actin alpha 1	Acta1	377	42.1	70.82
P62737	肌动蛋白α2 Actin alpha 2	Acta2	377	42.0	72.41
P63260	肌动蛋白γ Actin, gamma	Actg1	375	41.8	73.60
Q9R0Q6	肌动蛋白相关蛋白2/3复合体,1A亚基	Arpc1a	370	41.6	8.38
	Actin related protein 2/3 complex, subunit 1A
P11276	纤连蛋白1 Ffibronectin1	Fn1	2 477	272.5	2.10
Q9JKF1	IQ基序包含GTP酶激活蛋白1	Iqgap1	1 657	188.7	3.98
	IQ motif containing GTPase activating protein 1
Q9R0U0	富含丝氨酸和精氨酸的剪接因子10	Srsf10	262	31.3	4.20
	Serine and arginine-rich splicing factor 10
Q8BL97	富含丝氨酸和精氨酸的剪接因子7	Srsf7	267	30.8	5.24
	Serine and arginine-rich splicing factor 7
Q8CI51	PDZ和LIM结构域5 PDZ and LIM domain 5	Pdlim5	591	63.3	3.22
Q9CZD3	甘氨酸-tRNA合成酶Glycyl-tRNA synthetase	Gars	729	81.9	1.78
Q99JI6	RAS相关蛋白1b RAS related protein 1b	Rap1b	184	20.8	6.52
P54822	腺苷酸基琥珀酸裂解酶Adenylosuccinate lyase	Adsl	484	54.9	15.29

UniProt登录号	蛋白质名称	基因名	长度	分子量	覆盖度
UniProt accession	Protein name	Gene name	Length/aa	Mass/ku	Coverage/%
P97461	核糖体蛋白S5 Ribosomal protein S5	Rps5	204	22.9	4.41
Q9CZX8	核糖体蛋白S19 Ribosomal protein S19	Rps19	145	16.1	8.97
P60867	核糖体蛋白S20 Ribosomal protein S20	Rps20	119	13.4	10.08
P62858	核糖体蛋白S28 Ribosomal protein S28	Rps28	69	7.8	17.39
P62274	核糖体蛋白S29 Ribosomal protein S29	Rps29	56	6.7	14.29
Q9D8E6	核糖体蛋白L4 Ribosomal protein L4	Rpl4	419	47.2	12.89
P47962	核糖体蛋白L5 Ribosomal protein L5	Rpl5	297	34.4	12.79
O09167	核糖体蛋白L21 Ribosomal protein L21	Rpl21	160	18.6	6.88
P61255	核糖体蛋白L26 Ribosomal protein L26	Rpl26	145	17.3	9.66
P62900	核糖体蛋白L31 Ribosomal protein L31	Rpl31	125	14.5	18.40
P62717	核糖体蛋白L18A Ribosomal protein L18A	Rpl18a	176	20.7	15.34
P99027	核糖体蛋白,大P2 Ribosomal protein, large P2	Rplp2	115	11.7	25.22
Q7TPR4	肌动蛋白1 Actinin 1	Actn1	892	103.1	4.04
P68134	肌动蛋白α1 Actin alpha 1	Acta1	377	42.1	70.82
P62737	肌动蛋白α2 Actin alpha 2	Acta2	377	42.0	72.41
P63260	肌动蛋白γ Actin, gamma	Actg1	375	41.8	73.60
Q9R0Q6	肌动蛋白相关蛋白2/3复合体,1A亚基	Arpc1a	370	41.6	8.38
	Actin related protein 2/3 complex, subunit 1A
P11276	纤连蛋白1 Ffibronectin1	Fn1	2 477	272.5	2.10
Q9JKF1	IQ基序包含GTP酶激活蛋白1	Iqgap1	1 657	188.7	3.98
	IQ motif containing GTPase activating protein 1
Q9R0U0	富含丝氨酸和精氨酸的剪接因子10	Srsf10	262	31.3	4.20
	Serine and arginine-rich splicing factor 10
Q8BL97	富含丝氨酸和精氨酸的剪接因子7	Srsf7	267	30.8	5.24
	Serine and arginine-rich splicing factor 7
Q8CI51	PDZ和LIM结构域5 PDZ and LIM domain 5	Pdlim5	591	63.3	3.22
Q9CZD3	甘氨酸-tRNA合成酶Glycyl-tRNA synthetase	Gars	729	81.9	1.78
Q99JI6	RAS相关蛋白1b RAS related protein 1b	Rap1b	184	20.8	6.52
P54822	腺苷酸基琥珀酸裂解酶Adenylosuccinate lyase	Adsl	484	54.9	15.29

Screening and functional analysis of proteins interacting with chicken adenylosuccinate lyase

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