Human AZFb deletions cause distinct testicular pathologies depending on their extensions in Yq11 and the Y haplogroup: new cases and review of literature

Abstract

Genomic AZFb deletions in Yq11 coined "classical" (i.e. length of Y DNA deletion: 6.23 Mb) are associated with meiotic arrest (MA) of patient spermatogenesis, i.e., absence of any postmeiotic germ cells. These AZFb deletions are caused by non-allelic homologous recombination (NAHR) events between identical sequence blocks located in the proximal arm of the P5 palindrome and within P1.2, a 92 kb long sequence block located in the P1 palindrome structure of AZFc in Yq11. This large genomic Y region includes deletion of 6 protein encoding Y genes, EIFA1Y, HSFY, PRY, RBMY1, RPS4Y, SMCY. Additionally, one copy of CDY2 and XKRY located in the proximal P5 palindrome and one copy of BPY1, two copies of DAZ located in the P2 palindrome, and one copy of CDY1 located proximal to P1.2 are included within this AZFb microdeletion. It overlaps thus distally along 2.3 Mb with the proximal part of the genomic AZFc deletion. However, AZFb deletions have been also reported with distinct break sites in the proximal and/or distal AZFb breakpoint intervals on the Y chromosome of infertile men. These so called "non-classical" AZFb deletions are associated with variable testicular pathologies, including meiotic arrest, cryptozoospermia, severe oligozoospermia, or oligoasthenoteratozoospermia (OAT syndrome), respectively. This raised the question whether there are any specific length(s) of the AZFb deletion interval along Yq11 required to cause meiotic arrest of the patient's spermatogenesis, respectively, whether there is any single AZFb Y gene deletion also able to cause this "classical" AZFb testicular pathology? Review of the literature and more cases with "classical" and "non-classical" AZFb deletions analysed in our lab since the last 20 years suggests that the composition of the genomic Y sequence in AZFb is variable in men with distinct Y haplogroups especially in the distal AZFb region overlapping with the proximal AZFc deletion interval and that its extension can be "polymorphic" in the P3 palindrome. That means this AZFb subinterval can be rearranged or deleted also on the Y chromosome of fertile men. Any AZFb deletion observed in infertile men with azoospermia should therefore be confirmed as "de novo" mutation event, i.e., not present on the Y chromosome of the patient's father or fertile brother before it is considered as causative agent for man's infertility. Moreover, its molecular length in Yq11 should be comparable to that of the "classical" AZFb deletion, before meiotic arrest is prognosed as the patient's testicular pathology.

Keywords: AZFb Y genes; AZFb “de novo” and “polymorphic” deletions; AZFb-c amplicons; Meiotic arrest; OAT syndrome; Y-haplogroups; “Classical” and “non-classical” AZFb deletions.

Fig. 1

Schematic view on euchromatic part of the long arm of the human Y chromosome with focus on Yq11.22 including the “classical” AZFb deletion interval also known as (P5/proximal P1.2 deletion) encompassing 6.23 Mb genomic Y DNA and overlapping with AZFc for 2.3 Mb in Yq11.23 (for more details see ref [12]). a Repetitive sequence blocks with high sequence homology (< 99.7%) and designated as “amplicons” are displayed with a specific colour code as follows: yel(low) 1, 2, 3; b(lue) 1, 2, 3, 5, 6, t(urquoise) 1, 2; g(reen) 1; r(ed) 1, 2; gr(ey) 1. They are organized in 5 palindromes (P1–P5) as indicated. The X homologous single copy regions in AZFb are marked with: u(nique) 1. u(nique) 2,3 are Y specific single copy spacer regions of amplicons. DYZ19 is a tandem repetitive sequence block composed with 125 nts long sequence units along 400 kb (i.e. 3200 repeats). STS markers used in the clinic for diagnostic analyses of AZFb deletions are given with their positions above the amplicon structure. Those coloured in orange (+STS present; −STS absent) are used in ref. [16] to curtail the putative AZFb break sites of a “classical” AZFb deletion according to ref. [12] and including deletion of all Y genes listed in b. STS markers coloured in grey are used in ref. [17]. Absence of neighboured sY127 and sY134 in u1 is used in ref. [17] to indicate presence of complete AZFb deletions. b Schematic map of positions and polarities of the Y genes in AZFb encoding proteins according to ref. [11, 12]. RBMY1 is composed of 6 gene copies located in two distinct Y regions from proximal to distal: RBMY1B, RBMY1A1, RBMY1D, RBMY1E, in u1 and RBMY1F, RBMY1J, in t1 and t2, respectively. All AZFb Y genes are expressed in male germ cells during spermatogenesis as described in the main text

Fig. 2

Schematic view on extension of AZFb deletion in Yq11 of an infertile man with OAT syndrome (case ID: AZG189). a PCR multiplex assays used for analyses of AZFb Y gene deletions [16] indicated a partial AZFb deletion because the “classical” proximal and distal AZFb breakpoint regions were still present (sY1227: + ; sY1291: +). b Comparison of PCR amplification product intensities for PRY on the Y chromosome of male control (man with normal Y chromosome, 2 PRY genes, and normal fertility) and of AZG189 in B mix assay reveals that only one PRY gene copy is deleted. c Molecular sequence analyses of AZFb break-fusion site on the AZFG189 Y chromosome revealed non allelic homologous recombination event (NAHR) between two T_n tracts located in the spacer region of the two b5 amplicons and in the t1 amplicon distal to the first PRY gene copy. Molecular length of this AZFb deletion is 4.45 Mb. Using the genomic STS markers, sY105, sY121, sY127, sY134, sY143, sY153 (grey coloured in a) recommended by the EMQN/EAA best practice guidelines [17] this AZFb deletion would have been diagnosed as “classical” AZFb deletion. For further discussion see main-text

Fig. 3

a Schematic view on extensive map of genomic STS markers flanking the AZFb Y genes in the AZFb interval of the human Y chromosome to distinguish the extension of partial AZFb deletions of patients with distinct testicular pathologies. b The SNV deletion map of these patients for the distal AZFb-AZFc overlap region at the right is presented in Table 2 and therefore only partly viewed here in this scheme

Fig. 4

Immunohistochemical staining of human testis tissue sections with specific CREM antibodies marks predominant expression of CREM in postmeiotic spermatids as described in ref. [42]. a CREM expression in spermatids (spd; see arrow) of testicular tissue section of man with normal fertility and b in spermatids of testicular tissue section of AZFG620 with “non classical” AZFb deletion including all AZFb Y genes. Scale bars: 50 µm